How Different Dietary Methionine Sources Could Modulate the Hepatic Metabolism in Rainbow Trout?

In aquafeeds in which plant proteins are used to replace fishmeal, exogenous methionine (Met) sources are demanded to balance the amino acid composition of diets and meet the metabolic fish requirements. Nonetheless, since different synthetic Met sources are commercially available, it is important to determine their bioavailability and efficacy. To address this issue, we conducted a two-month feeding trial with rainbow trout (Oncorhynchus mykiss), which were fed diets supplemented with five different forms of Met: Met-Met, L-Met, HMTBa, DL-Met, and Co DL-Met. No differences in growth performance were found in trout fed with different Met forms, but changes in the whole-body composition were found. In particular, Met-Met and L-Met promoted a significant body lipid reduction, whereas the protein retention was significantly increased in fish fed with HMTBa and Co DL-Met. The latter affected the hepatic Met metabolism promoting the trans-sulfuration pathway through the upregulation of CBS gene expression. Similarly, the L-Met enhanced the remethylation pathway through an increase in BHMT gene expression to maintain the cellular demand for Met. Altogether, our findings suggest an optimal dietary intake of all tested Met sources with similar promoting effects on fish growth and hepatic Met metabolism. Nevertheless, the mechanisms underlying these effects warrant further investigation.

Oxygen for the Newborn: Friend or Foe?

Oxygen supplementation is widely used in neonatal care, however, it can also cause toxic effects if not used properly. Therefore, it appears crucial to find a balance in oxygen administration to avoid damage as a consequence of its insufficient or excessive use. Oxygen toxicity is mainly due to the production of oxygen radicals, molecules normally produced in humans and involved in a myriad of physiological reactions. In the neonatal period, an imbalance between oxidants and antioxidant defenses, the so-called oxidative stress, might occur, causing severe pathological consequences. In this review, we focus on the mechanisms of the production of oxygen radicals and their physiological functions in determining a set of diseases grouped together as "free radical diseases in the neonate". In addition, we describe the evolution of the oxygenation target recommendations during neonatal resuscitation and post-stabilization phases with the aim to define the best oxygen administration according to the newest evidence.

Glycemic Control, Basal/Bolus Distribution, BMI and Meal Management in Children and Adolescents with Type 1 Diabetes and Advanced Hybrid Closed Loop.

Evidence about the impact of advanced hybrid closed loop (AHCL) on body mass index (BMI) and eating habits in children with type 1 diabetes (T1D) is lacking. This real-world study aimed at evaluating glycemic control, BMI, meals and basal/bolus distribution in young subjects with T1D treated by AHCL. Glycemic metrics, HbA1c, basal/bolus distribution, meals/day, BMI, total daily dose (TDD), and carbohydrates/kg (CHO/kg) have been evaluated in 83 subjects, aged 13 ± 4.5 years, in manual mode, 3 and 6 months after auto-mode. Time in range (TIR) increased after 3 months, exceeding the target of 70% and was maintained at 6 months. While coefficient of variation (CV) did not change, the glucose management indicator (GMI) decreased in auto-mode (6.7 ± 0.3 vs. 7.1 ± 0.5%; p < 0.001), as well as HbA1c. Basal proportion decreased in favor of boluses (38.3 ± 7.3 vs. 43.6 ± 10.9%; p < 0.001). Meals increased at 3 and 6 months (4.4 ± 1.2 vs. 5.0 ± 1.5, p 0.002 and 5.1 ± 1.7, p < 0.001), as well as TDD/kg, without changes in BMI and CHO consumed. No differences in meal composition have arisen from food diaries. In conclusion, AHCL ensured the achievement and maintenance of target TIR in young T1D subjects. The number of meals, TDD, and insulin bolus proportion increased over time, but BMI remained stable.

New value from food and industrial wastes - Bioaccumulation of omega-3 fatty acids from an oleaginous microbial biomass paired with a brewery by-product using black soldier fly (Hermetia illucens) larvae.

Research on bioconversion based on insects is intensifying as it addresses the problem of reducing and reusing food and industrial waste. To reach this goal, we need to find more means of pairing waste to insects. With this goal, brewers' spent grains (BSG) - a food waste of the brewing industry - paired with the oleaginous biomass of the thraustochytrid Schizochytrium limacinum cultivated on crude glycerol - a major waste of biodiesel production - were successfully used to grow Hermetia illucens larvae. Combining BSG and S. limacinum in the diet in an attempt to design the lipid profile of H. illucens larvae to contain a higher percentage of omega-3 fatty acids is novel. Insect larvae were grown on three different substrates: i) standard diet for Diptera (SD), ii) BSG, and iii) BSG + 10% S. limacinum biomass. The larvae and substrates were analyzed for fatty acid composition and larval growth was measured until 25% of insects reached the prepupal stage. Our data showed that including omega-3-rich S. limacinum biomass in the BSG substrate promoted an increase in larval weight compared to larvae fed on SD or BSG substrates. Furthermore, it was possible, albeit in a limited way, to incorporate omega-3 fatty acids, principally docosahexaenoic acid (DHA) from BSG + S. limacinum substrate containing 20% of DHA into the larval fat (7% DHA). However, H. illucens with this level of DHA may not be suitable if the aim is to get larvae with high omega-3 lipids to feed carnivorous fish.

Effect of partial substitution of fishmeal with insect meal (Hermetia illucens) on gut neuromuscular function in Gilthead sea bream (Sparus aurata).

Alternative nutrient sources to fishmeal for fish feed, such as insect meals, represent a promising sustainable supply. However, the consequences for fish digestive function have not been exhaustively investigated. In the present study we evaluated the effect of partial fishmeal substitution with 10% Hermetia illucens (Hi10) larvae meal on the neuromuscular function of proximal and distal intestine in gilthead sea bream. In animals fed with insect meal, weight and growth parameters were similar to controls fed with conventional fishmeal. In addition, no anomalies in intestinal gross morphology and no overt signs of inflammation were observed. The gastrointestinal transit was significantly reduced in Hi10 fed animals. In the proximal and distal intestine longitudinal muscle, Hi10 feeding downregulated the excitatory cholinergic and serotoninergic transmission. Sodium nitroprusside-induced inhibitory relaxations increased in the proximal intestine and decreased in the distal intestine after Hi10 meal. Changes in the excitatory and inhibitory components of peristalsis were associated with adaptive changes in the chemical coding of both proximal and distal intestine myenteric plexus. However, these neuromuscular function alterations were not associated with considerable variations in morphometric growth parameters, suggesting that 10% Hi meal may represent a tolerable alternative protein source for gilthead sea bream diets.

Effects of Partially Defatted Hermetia illucens Meal in Rainbow Trout Diet on Hepatic Methionine Metabolism.

This study investigated, for the first time, the effects of replacement of fishmeal (FM) with insect meal from Hermetia illucens (HI) on the transcript levels of three genes involved in methionine (Met) metabolism in rainbow trout (Oncorhynchus mykiss) liver. Two target genes-betaine-homocysteine S-methyltransferase (BHMT) and S-adenosylhomocysteine hydrolase (SAHH)-are involved in Met resynthesis and the third one-cystathionine ß synthase (CBS)-is involved in net Met loss (taurine synthesis). We also investigated the levels of two Met metabolites involved in the maintenance of methyl groups and homocysteine homeostasis in the hepatic tissue: S-adenosylmethionine (SAM) and S-adenosylhomocysteine (SAH). Three diets were formulated, an FM-based diet (HI0) and two diets in which 25% (HI25) and 50% (HI50) of FM was replaced with HI larvae meal. A 78-day feeding trial involved 360 rainbow trout with 178.9 ± 9.81 g initial average weight. Dietary replacement of up to 50% of FM with HI larvae meal, without any Met supplementation, did not negatively affect rainbow trout growth parameters and hepatic Met metabolism. In particular, Met availability from the insect-based diets directly modulated the transcript levels of two out of three target genes (CBS, SAHH) to maintain an optimal level of one-carbon metabolic substrates, i.e., the SAM:SAH ratio in the hepatic tissue.

Effect of dietary oil from Camelina sativa on the growth performance, fillet fatty acid profile and gut microbiome of gilthead Sea bream (Sparus aurata).

BACKGROUND: In the last two decades, research has focused on testing cheaper and sustainable alternatives to fish oil (FO), such as vegetable oils (VO), in aquafeeds. However, FO cannot be entirely replaced by VOs due to their lack of omega-3 (n-3) long-chain polyunsaturated fatty acids (LC-PUFA), particularly eicosapentaenoic (EPA; 20:5n-3) and docosahexaenoic (DHA; 22:6n-3) acids. The oilseed plant, Camelina sativa, may have a higher potential to replace FO since it can contains up to 40% of the omega-3 precursors α-linolenic acid (ALA; 18:3n-3) and linoleic acid (LA; 18:2n-6). METHODS: A 90-day feeding trial was conducted with 600 gilthead sea bream (Sparus aurata) of 32.92 ±  0.31 g mean initial weight fed three diets that replaced 20%, 40% and 60% of FO with CO and a control diet of FO. Fish were distributed into triplicate tanks per diet and with 50 fish each in a flow-through open marine system. Growth performance and fatty acid profiles of the fillet were analysed. The Illumina MiSeq platform for sequencing of 16S rRNA gene and Mothur pipeline were used to identify bacteria in the faeces, gut mucosa and diets in addition to metagenomic analysis by PICRUSt. RESULTS AND CONCLUSIONS: The feed conversion rate and specific growth rate were not affected by diet, although final weight was significantly lower for fish fed the 60% CO diet. Reduced final weight was attributed to lower levels of EPA and DHA in the CO ingredient. The lipid profile of fillets were similar between the dietary groups in regards to total saturated, monounsaturated, PUFA (n-3 and n-6), and the ratio of n-3/n-6. Levels of EPA and DHA in the fillet reflected the progressive replacement of FO by CO in the diet and the EPA was significantly lower in fish fed the 60% CO diet, while ALA was increased. Alpha and beta-diversities of gut bacteria in both the faeces and mucosa were not affected by any dietary treatment, although a few indicator bacteria, such as Corynebacterium and Rhodospirillales, were associated with the 60% CO diet. However, lower abundance of lactic acid bacteria, specifically Lactobacillus, in the gut of fish fed the 60% CO diet may indicate a potential negative effect on gut microbiota. PICRUSt analysis revealed similar predictive functions of bacteria in the faeces and mucosa, although a higher abundance of Corynebacterium in the mucosa of fish fed 60% CO diet increased the KEGG pathway of fatty acid synthesis and may act to compensate for the lack of fatty acids in the diet. In summary, this study demonstrated that up to 40% of FO can be replaced with CO without negative effects on growth performance, fillet composition and gut microbiota of gilthead sea bream.

Protective Effect of Dietary Taurine from ROS Production in European Seabass under Conditions of Forced Swimming.

Taurine (Tau) is an amino sulfonic acid, which is widely distributed in animal tissues, whereas it is almost lacking in plants with the exception of certain algae, seaweeds, and few others. In the aquafeed industry, Tau is mainly used as a feed additive to promote growth in marine fish species with limited cysteine sulfinate decarboxylase activity. In particular, Tau supplementation is required in feeds in which fishmeal (FM) is substituted with high percentages of plant-derived protein sources such as soybean meals (SBM) that have much lower levels of Tau than FM. In addition to being a growth promoter, Tau exert powerful antioxidant properties being a scavenger of the reactive oxygen species (ROS). Under sustained swimming conditions, an intracellular increase in ROS production can occur in fish red muscle where the abundance of mitochondria (the main site of ROS formation) is high. Accordingly, this study aimed at investigating the effects of dietary Tau on European seabass (Dicentrarchus labrax) growth and oxidative stress response induced by swimming exercise. Individually tagged fish of 92.57 ± 20.33 g mean initial weight were fed two experimental diets containing the same low percentage of FM and high percentage of SBM. One diet was supplemented with 1.5% of Tau. Tau supplemented in the diet had a positive effect on fish growth, and enhanced swimming performance and antioxidant status. Two swim endurance tests were performed during the feeding trial. Metabolic oxygen consumption (MO2) was measured during exercise at incremental swimming speeds (0.7, 1.4, 2.1, 2.8, 3.5, and then 4.2 BL (body length) s-1, until fatigue). Fish maximal sustainable swimming speed (Ucrit) was determined too. To investigate the antioxidant effect of dietary Tau, we also measured ROS production in fish blood by RBA (respiratory burst activity) assay and quantified the expression of genes coding for antioxidant enzymes by qPCR (quantitative polymerase chain reaction) , such as SOD (superoxide dismutase), GPX (glutathione peroxidase), and CAT (catalase) in red muscle and liver. There was a significant effect of Tau upon Ucrit during exercise. Additionally, ROS production was significantly lower in fish fed with Tau supplemented diet, supporting the role of Tau as ROS scavenger. The protective effect of Tau against oxidative stress induced by forced swimming was denoted also by a significant decrease in antioxidant enzymes gene expression in fish liver and muscle. Taken together these results demonstrate that Tau is beneficial in low FM-based diets for seabass.

Neurochemical characterization of myenteric neurons in the juvenile gilthead sea bream (Sparus aurata) intestine.

We evaluated the chemical coding of the myenteric plexus in the proximal and distal intestine of gilthead sea bream (Sparus aurata), which represents one of the most farmed fish in the Mediterranean area. The presence of nitric oxide (NO), acetylcholine (ACh), serotonin (5-HT), calcitonin-gene-related peptide (CGRP), substance P (SP) and vasoactive intestinal peptide (VIP) containing neurons, was investigated in intestinal whole mount preparations of the longitudinal muscle with attached the myenteric plexus (LMMP) by means of immunohistochemical fluorescence staining. The main excitatory and inhibitory neurochemicals identified in intestinal smooth muscle were ACh, SP, 5HT, and NO, VIP, CGRP. Some neurons displayed morphological features of ascending and descending interneurons and of putative sensory neurons. The expression of these pathways in the two intestinal regions is largely superimposable, although some differences emerged, which may be relevant to the morphological properties of each region. The most important variances are the higher neuronal density and soma size in the proximal intestine, which may depend on the volume of the target tissue. Since in the fish gut the submucosal plexus is less developed, myenteric neurons substantially innervate also the submucosal and epithelial layers, which display a major thickness and surface in the proximal intestine. In addition, myenteric neurons containing ACh and SP, which mainly represent excitatory motor neurons and interneurons innervating the smooth muscle were more numerous in the distal intestine, possibly to sustain motility in the thicker smooth muscle coat. Overall, this study expands our knowledge of the intrinsic innervation that regulates intestinal secretion, absorption and motility in gilthead sea bream and provides useful background information for rational design of functional feeds aimed at improving fish gut health.

Effects of Sodium Butyrate Treatment on Histone Modifications and the Expression of Genes Related to Epigenetic Regulatory Mechanisms and Immune Response in European Sea Bass (Dicentrarchus Labrax) Fed a Plant-Based Diet.

Bacteria that inhabit the epithelium of the animals' digestive tract provide the essential biochemical pathways for fermenting otherwise indigestible dietary fibers, leading to the production of short-chain fatty acids (SCFAs). Of the major SCFAs, butyrate has received particular attention due to its numerous positive effects on the health of the intestinal tract and peripheral tissues. The mechanisms of action of this four-carbon chain organic acid are different; many of these are related to its potent regulatory effect on gene expression since butyrate is a histone deacetylase inhibitor that play a predominant role in the epigenetic regulation of gene expression and cell function. In the present work, we investigated in the European sea bass (Dicentrarchus labrax) the effects of butyrate used as a feed additive on fish epigenetics as well as its regulatory role in mucosal protection and immune homeostasis through impact on gene expression. Seven target genes related to inflammatory response and reinforcement of the epithelial defense barrier [tnfα (tumor necrosis factor alpha) il1ß, (interleukin 1beta), il-6, il-8, il-10, and muc2 (mucin 2)] and five target genes related to epigenetic modifications [dicer1(double-stranded RNA-specific endoribonuclease), ehmt2 (euchromatic histone-lysine-N-methyltransferase 2), pcgf2 (polycomb group ring finger 2), hdac11 (histone deacetylase-11), and jarid2a (jumonji)] were analyzed in fish intestine and liver. We also investigated the effect of dietary butyrate supplementation on histone acetylation, by performing an immunoblotting analysis on liver core histone extracts. Results of the eight-week-long feeding trial showed no significant differences in weight gain or SGR (specific growth rate) of sea bass that received 0.2% sodium butyrate supplementation in the diet in comparison to control fish that received a diet without Na-butyrate. Dietary butyrate led to a twofold increase in the acetylation level of histone H4 at lysine 8, but showed no effect on the histone H3 at Lys9. Moreover, two different isoforms of histone H3 that might correspond to the H3.1 and H3.2 isoforms previously found in terrestrial animals were separated on the immunoblots. The expression of four (il1 ß, il8, irf1, and tnfα) out of seven analyzed genes related to mucosal protection and inflammatory response was significantly different between the two analyzed tissues but only il10 showed differences in expression due to the interaction between tissue and butyrate treatment. In addition, butyrate caused significant changes in vivo in the expression of genes related to epigenetic regulatory mechanisms such as hdac11, ehmt2, and dicer1. Statistical analysis by two-way ANOVA for these genes showed not only significant differences due to the butyrate treatment, but also due to the interaction between tissue and treatment.