Hepatic Global DNA Hypomethylation Phenotype in Rainbow Trout Fed Diets Varying in Carbohydrate to Protein Ratio.

BACKGROUND: A high carbohydrate-low protein diet can induce hepatic global DNA hypomethylation in trout. The mechanisms remain unclear. OBJECTIVES: We aimed to investigate whether an increase in dietary carbohydrates (dHCs) or a decrease in dietary proteins (dLPs) can cause hepatic global DNA hypomethylation, as well as explore the underlying mechanisms in trout. METHODS: Two feeding trials were conducted on juvenile males, both of which involved a 4-d fasting and 4-d refeeding protocol. In trial 1, trout were fed either a high protein-no carbohydrate [HP-NC, protein 60% dry matter (DM), carbohydrates 0% DM] or a moderate protein-high carbohydrate (MP-HC, protein 40% DM, carbohydrates 30% DM) diet. In trial 2, fish were fed either a moderate protein-no carbohydrate (MP-NC, protein 40% DM, carbohydrates 0% DM), an MP-HC (protein 40% DM, carbohydrates 30% DM), or a low protein-no carbohydrate (LP-NC, protein 20% DM, carbohydrates 0% DM) diet to separate the effects of dHCs and dLPs on the hepatic methylome. Global CmCGG methylation, DNA demethylation derivative concentrations, and mRNA expression of DNA (de)methylation-related genes were measured. Differences were tested by 1-factor ANOVA when data were normally distributed or by Kruskal-Wallis nonparametric test if not. RESULTS: In both trials, global CmCGG methylation concentrations remained unaffected, but the hepatic 5-mdC content decreased after refeeding (1-3%). The MP-HC group had 3.4-fold higher hepatic 5-hmdC and a similar 5-mdC concentration compared with the HP-NC group in trial 1. Both MP-HC and LP-NC diets lowered the hepatic 5-mdC content (1-2%), but only the LP-NC group had a significantly lower 5-hmdC concentration (P < 0.01) compared with MP-NC group in trial 2. CONCLUSIONS: dHC and dLP independently induced hepatic global DNA demethylation in trout. The alterations in other methylation derivative concentrations indicated the demethylation process was achieved through an active demethylation pathway and probably occurred at non-CmCGG sites.

Why Do Some Rainbow Trout Genotypes Grow Better With a Complete Plant-Based Diet? Transcriptomic and Physiological Analyses on Three Isogenic Lines.

Within the context of a growing aquaculture production coupled with a plateau of the production in the main components of aquafeeds (fish oil and fishmeal), recent studies have typically focused on replacing these feedstuffs with terrestrial plant ingredients for cultured carnivorous aquatic species, such as rainbow trout (Oncorhynchus mykiss). Substitution rates without adverse effects have, however, reached their limit. One potential way forward would be to take advantage of the genetic variability that exists in the salmonid population. However, to date, little is known about the underlying molecular mechanisms responsible for this genetic variability. The aim of the present research was to understand why some genotypes are better able to utilize plant-based diets devoid of marine resources. In this regard, three isogenic lines of rainbow trout (R23h, AB1h, and A22h), with similar growth when fed marine resources-based diets and which differ greatly in their responses to a plant-based diet, were fed with either a complete plant-based diet (V diet) or a marine resources-based diet (M diet) since first-feeding. Fish traits and the hepatic transcriptome of these three genotypes were compared after 5 months of feeding. First, differences in the ability to grow with the V diet observed amongst genotypes was not due to higher feed intake, but instead due to differences in feed efficiency. The comparison of transcriptome profiles revealed 575 (R23h vs. AB1h), 1,770 (R23h vs. A22h), and 2,973 (AB1h vs. A22h) probes differentially expressed amongst the three genotypes when fed the V diet. Interestingly, R23h and AB1h fish, which were the least affected by the V diet, exhibited the highest growth. These results demonstrate that these fish were able to maintain a high level of energy production and protein synthesis. Moreover, these genotypes were also able to activate pathways linked to lipid and cholesterol metabolisms, such as the biosynthesis of long-chain polyunsaturated fatty acids. Finally, as previously, immunity seems to also play an important role in the ability of fish to use the V diet, and further studies are needed to understand the mechanisms by which immunity interacts with growth.

Impact of feed restriction and fragmented feed distribution on performance, intake behaviour and digestion of the growing rabbit.

Postweaning feed restriction preserves rabbit digestive health after weaning, but the underlying physiological mechanisms are not yet understood. To elucidate whether the feeding intake pattern modification related to feed restriction might be involved, we studied the effects of both feed intake quantity and intake frequency. Animals were allotted at weaning (28 d old) in a 2 × 2 factorial design: feed intake quantity (AL = ad libitum vs R = 75% of AL) and fragmented feed distribution (FFD) (1 vs 13 distributions), thus forming four groups (AL1, AL13, R1 and R13). New Zealand White growing rabbits were used from weaning to slaughter (70 d old), to analyse mortality, morbidity, performance, intake behaviour, digestion and microbial activity. Seven days after starting feed restriction (35 d old, group R1), rabbits consumed 44% of the feed within 2 h, 65% in 4 h and in 7 h over 95%. Over the 28-70 d period, mortality was low (5.3%) while morbidity averaged 18.5% and neither was affected by treatment. However, FFD tended to decrease the morbidity rate during the first 14 days after weaning (P = 0.06). Feed conversion (28-70 d) was improved by restriction (+15%, P < 0.001) and by FFD (+5%, P < 0.001). Nutrient digestibility was improved by restriction (+10% for energy, P < 0.01), but not by FFD. Fragmented feed distribution led to a lower stomachal pH, in the antrum (1.48 vs 2.13, P < 0.001) and in the fundus (1.52 vs 2.63, P < 0.001), while a higher pH was found in the caecum (6.07 vs 5.86, P < 0.001). Butyrate proportion in the caecum was reduced by four units for restricted groups. Fragmented feed distribution reduced the caecal VFA concentration by 23% within restricted rabbit groups only. A similar interaction between intake level and FFD was observed for fibrolytic activity (cellulase and xylanase). The diversity of caecal bacterial community was not modified by either of the two factors studied. Globally, fragmented meals have no major impacts on the caecal microbial activity, diversity, and thus would not be implicated in the better resistance of restricted rabbit to digestive troubles.

Exploring the Impact of a Low-Protein High-Carbohydrate Diet in Mature Broodstock of a Glucose-Intolerant Teleost, the Rainbow Trout.

Sustainable aquaculture production requires a greater reduction in the use of marine-derived ingredients, and one of the most promising solutions today is the augmentation in the proportion of digestible carbohydrates in aquafeed. This challenge is particularly difficult for high trophic level teleost fish as they are considered to be glucose-intolerant (growth delay and persistent postprandial hyperglycemia observed in juveniles fed a diet containing more than 20% of carbohydrates). It was previously suggested that broodstock could potentially use carbohydrates more efficiently than juveniles, probably due to important metabolic changes that occur during gametogenesis. To investigate this hypothesis, 2-year old male and female rainbow trout (Oncorhynchus mykiss) were either fed a diet containing no carbohydrates (NC) or a 35%-carbohydrate diet (HC) for an entire reproductive cycle. Zootechnical parameters as well as the activities of enzymes involved in carbohydrate metabolism were measured in livers and gonads. Fish were then reproduced to investigate the effects of such a diet on reproductive performance. Broodstock consumed the HC diet, and in contrast to what is commonly observed in juveniles, they were able to grow normally and they did not display postprandial hyperglycemia. The modulation of their hepatic metabolism, with an augmentation of the glycogenesis, the pentose phosphate pathway and a possible better regulation of gluconeogenesis, may explain their improved ability to use dietary carbohydrates. Although the HC diet did induce precocious maturation, the reproductive performance of fish was not affected, confirming that broodstock are able to reproduce when fed a low-protein high-carbohydrate diet. In conclusion, this exploratory work has shown that broodstock are able to use a diet containing digestible carbohydrates as high as 35% and can then grow and reproduce normally over an entire reproductive cycle for females and at least at the beginning of the cycle for males. These results are highly promising and suggest that dietary carbohydrates can at least partially replace proteins in broodstock aquafeed.

Rainbow trout prefer diets rich in omega-3 long chain polyunsaturated fatty acids DHA and EPA.

The control of feed intake in fish in aquaculture requires the development of new techniques to improve diet composition, feed conversion efficiency and growth. The aim must be sustainability and an effective use of resources. The effect of replacing traditional aqua-feed ingredients (fishmeal and fish oil) by a 100% plant-based diet is known to drastically decrease fish performance (survival and growth). The present study examined the feed preference of rainbow trout Oncorhynchus mykiss for three diets containing distinct levels of omega-3 long chain polyunsaturated fatty acids (ω-3 LCPUFA): eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) (0% for low, 5% for medium and 20% for high, total fatty acid content). Feed preference values for each group (low v. medium ω-3 diets, medium v. high ω-3 diets and low v. high ω-3 diets) were observed using two self-feeders positioned at opposite sides of the tank. The hypothesis was that the decrease of fish growth and survival rate of fish fed with 100% plant-based diet could be explained by the absence of ω-3 LCPUFA relating to decrease of food intake. This could explain the tasting role of ω-3 LCPUFA in the feeding behavior of rainbow trout (which reflects the motivation to consume feed). The results showed that rainbow trout could discriminate between the diets containing different level of ω-3 LCPUFA even if unable to differentiate between level of 5% (no preference observed in low v. medium ω-3 diets). Overall they had a preference for diet high in ω-3 LCPUFA: 59.5% preference for high ω-3 diet in high v. low ω-3 diets, and 75.6% preference for high ω-3 diet in medium v. high ω-3 diets respectively. This preference was repeated after 21 days and for a further 21 days when the feeds were exchanged between the two self- feeders in each tank: 63.3% preference for high ω-3 diet in high v. low ω-3 diets, and 69,5% preference for high ω-3 diet in medium v. high ω-3 diets respectively. The tests also indicated a difference in the extent of food waste of each of the three diets revealed by uneaten pellets after feed demands. During two periods of test, high ω-3 diet was the most appreciated, the least wasted and the most eaten (all choice groups) whereas the most uneaten feed remained the least appreciated diet in three choices diets (low ω-3 diet in low v. medium ω-3 diets, medium in medium v. high ω-3 diets and low in low v. high ω-3). In conclusion, this study highlighted the influence of ω-3 LCPUFA in the feeding behavior of juvenile rainbow trout, levels of ω-3 LCPUFA drove dietary choices in the fish.

Postprandial kinetics of gene expression of proteins involved in the digestive process in rainbow trout (O. mykiss) and impact of diet composition.

The impact of increased incorporation of plant ingredients on diets for rainbow trout was evaluated in terms of gene expression of gastric (gastric lipase, pepsinogen) and intestinal (prolidase, maltase, phospholipase A2) digestive enzymes and nutrient transporters (peptide and glucose transporters), as well as of postprandial levels of plasma glucose, triglycerides and total free amino acids. For that purpose, trout alevins were fed from the start of exogenous feeding one of three different experimental diets: a diet rich in fish meal and fish oil (FM-FO), a plant-based diet (noFM-noFO) totally free from fish meal and fish oil, but containing plant ingredients and a Mixed diet (Mixed) intermediate between the FM-FO and noFM-noFO diets. After 16 months of rearing, all fish were left unfed for 72 h and then given a single meal to satiation. Blood, stomach and anterior intestine were sampled before the meal and at 2, 6 and 12 h after this meal. The postprandial kinetics of gene expression of gastric and intestinal digestive enzymes and nutrient transporters were then followed in trout fed the FM-FO diet. The postprandial profiles showed that the expression of almost all genes studied was stimulated by the presence of nutrients in the digestive tract of trout, but the timing (appearance of peaks) varied between genes. Based on these data, we have focused on the molecular response to dietary factors in the stomach and the intestine at 6 and 12 h after feeding, respectively. The reduction in FM and FO levels of dietary incorporation induced a significant decrease in the gene expression of gastric lipase, GLUT2 and PEPT1. The plasma glucose and triglycerides levels were also reduced in trout fed the noFM-noFO diet. Consequently, the present study suggests a decrease in digestive capacities in trout fed a diet rich in plant ingredients.

Effect of low dose of fumonisins on pig health: immune status, intestinal microbiota and sensitivity to Salmonella.

The objective of this study was to measure the effects of chronic exposure to fumonisins via the ingestion of feed containing naturally contaminated corn in growing pigs infected or not with Salmonella spp. This exposure to a moderate dietary concentration of fumonisins (11.8 ppm) was sufficient to induce a biological effect in pigs (Sa/So ratio), but no mortality or pathology was observed over 63 days of exposure. No mortality or related clinical signs, even in cases of inoculation with Salmonella (5 × 104 CFU), were observed either. Fumonisins, at these concentrations, did not affect the ability of lymphocytes to proliferate in the presence of mitogens, but after seven days post-inoculation they led to inhibition of the ability of specific Salmonella lymphocytes to proliferate following exposure to a specific Salmonella antigen. However, the ingestion of fumonisins had no impact on Salmonella translocation or seroconversion in inoculated pigs. The inoculation of Salmonella did not affect faecal microbiota profiles, but exposure to moderate concentrations of fumonisins transiently affected the digestive microbiota balance. In cases of co-infection with fumonisins and Salmonella, the microbiota profiles were rapidly and clearly modified as early as 48 h post-Salmonella inoculation. Therefore under these experimental conditions, exposure to an average concentration of fumonisins in naturally contaminated feed had no effect on pig health but did affect the digestive microbiota balance, with Salmonella exposure amplifying this phenomenon.

Impact of deoxynivalenol on the intestinal microflora of pigs.

Deoxynivalenol (DON), a mycotoxin produced by some Fusarium species, is a frequent contaminant of cereal. In the present study, 24 weanling piglets received either control feed or feed naturally contaminated with DON (2.8 mg/kg) for four weeks. Consumption of contaminated feed significantly reduced the animal weight gain during the first week of the experiment, but had a moderate effect on cultivable bacteria in the pig intestine. By contrast, changes in the intestinal microflora were observed by Capillary Electrophoresis Single-Stranded Conformation Polymorphism (CE-SSCP) in DON-exposed animals, suggesting an impact of this toxin on the dynamics of intestinal bacteria communities.