Atlantic salmon (Salmo salar L.) parr fed genetically modified soybeans and maize: Histological, digestive, metabolic, and immunological investigations.

Physiological and health related responses to dietary inclusion of genetically modified (GM) full-fat soybean meal (Roundup Ready; GM-soy) and maize (MON810 Bt-maize; GM-maize), as well as non-parental, untransformed lines (nGM-soy and nGM-maize D2), were evaluated in farmed Atlantic salmon (Salmo salar L.) parr during the first 8 months of feeding. Significant effects of dietary GM presence were only found in intestinal Na+-dependent d-glucose uptake and SGLT1 protein level in the region pyloric caeca in which the highest values were found in the GM-soy, intermediate in the nGM-soy, and lowest in the standard FM fed groups. Data from this study confirm that GM soybeans (RRS) and maize (MON810) at inclusion levels of about 6% appear to be as safe as commercially available nGM soy and maize in diets for Atlantic salmon parr. Results from studies with higher inclusion levels and with non-modified, isogenic or near-isogenic parental lines as control groups are pending.

Evaluation of stress- and immune-response biomarkers in Atlantic salmon, Salmo salar L., fed different levels of genetically modified maize (Bt maize), compared with its near-isogenic parental line and a commercial suprex maize.

The present study was designed to evaluate if genetically modified (GM) maize (Bt maize, event MON810) compared with the near-isogenic non-modified (nGM) maize variety, added as a starch source at low or high inclusions, affected fish health of post-smolt Atlantic salmon, Salmo salar L. To evaluate the health impact, selected stress- and immune-response biomarkers were quantified at the gene transcript (mRNA) level, and some also at the protein level. The diets with low or high inclusions of GM maize, and its near-isogenic nGM parental line, were compared to a control diet containing GM-free suprex maize (reference diet) as the only starch source. Total superoxide dismutase (SOD) activity in liver and distal intestine was significantly higher in fish fed GM maize compared with fish fed nGM maize and with the reference diet group. Fish fed GM maize showed significantly lower catalase (CAT) activity in liver compared with fish fed nGM maize and to the reference diet group. In contrast, CAT activity in distal intestine was significantly higher for fish fed GM maize compared with fish fed reference diet. Protein level of heat shock protein 70 (HSP70) in liver was significantly higher in fish fed GM maize compared with fish fed the reference diet. No diet-related differences were found in normalized gene expression of SOD, CAT or HSP70 in liver or distal intestine. Normalized gene expression of interleukin-1 beta in spleen and head-kidney did not vary significantly between diet groups. Interestingly, fish fed high GM maize showed a significantly larger proportion of plasma granulocytes, a significantly larger sum of plasma granulocyte and monocyte proportions, but a significantly smaller proportion of plasma lymphocytes, compared with fish fed high nGM maize. In conclusion, Atlantic salmon fed GM maize showed some small changes in stress protein levels and activities, but none of these changes were comparable to the normalized gene expression levels analysed for these stress proteins. GM maize seemed to induce significant changes in white blood cell populations which are associated with an immune response.

Histological, digestive, metabolic, hormonal and some immune factor responses in Atlantic salmon, Salmo salar L., fed genetically modified soybeans.

The paper reports the second and final part of an experiment aiming to study physiological and health-related effects of genetically modified (GM) soybean meal (SBM) type Roundup Ready soybean (RRS) in diets for post-smolt Atlantic salmon. For 3 months salmon were fed diets containing 172 g kg(-1) full-fat SBM from RRS (GM-soy) or an unmodified, non-isogenic line (nGM-soy), or a reference diet with fishmeal as the sole protein source (FM). Slight differences in anti-nutrient levels were observed between the GM and nGM-soy. Histological changes were observed only in the distal intestine of the soy-fed fish. The incidence of moderate inflammation was higher in the GM-soy group (9 of 10 sampled fish) compared with the nGM-soy group (7 of 10). However, no differences in the concomitant decreases in activities of digestive enzymes located in the brush border (leucine aminopeptidase and maltase) and apical cytoplasm (acid phosphatase) of enterocytes or in the number of major histocompatibility complex class II+ cells, lysozyme activity, or total IgM of the distal intestine were observed. GM compared with nGM-soy fed fish had higher head kidney lysozyme (11,856 vs. 10,456 units g(-1) tissue) and a tendency towards higher acid phosphatase (0.45 vs. 0.39 micromol h(-1) kg(-1) body mass in whole tissue) activities, respectively. Plasma insulin and thyroxin levels, and hepatic fructose-1,6-bisphosphatase and ethoxyresorufin-O-deethylase activities were not significantly affected. It is not possible, however, to conclude whether the differences in responses to GM-soy were due to the genetic modification or to differences in soy cultivars in the soy-containing diets. Results from studies using non-modified, parental line soybeans as the control group are necessary to evaluate whether genetic modification of soybeans in diets poses any risk to farmed Atlantic salmon.

An examination of the intestinal tract of Atlantic salmon, Salmo salar L., parr fed different varieties of soy and maize.

This study was conducted to investigate the long-term effects of feeding plant products from both traditional breeding and from biotechnology on intestinal somatic indices, histology and cell proliferation in first-feeding Atlantic salmon, Salmo salar L. (initial weight 0.21 +/- 0.02 g). A standard fishmeal diet (standard fishmeal) was formulated to contain fishmeal as the sole protein source and suprex maize as the main starch source. Six experimental diets were then developed: two in which some of the fishmeal was replaced with commercially available, genetically modified Roundup Ready full-fat soybean meal (GM-soy) or commercially available, non-GM full-fat soybean meal (nGM-soy) at a level of 12.5% of the total diet, and four diets in which the suprex maize was replaced with two lines of GM-maize (Dekalb 1; D1 and Pioneer 1; P1), both products of event MON810, and their half-sibling non-GM counterparts (Dekalb 2; D2 and Pioneer 2; P2), at a level of 12.1% of total diet. Each diet was fed to fish in triplicate tanks and the experiment lasted for 8 months, during which the fish reached a final weight of 101-116 g. There was no significant effect of diet on the intestinal indices, nor were histological changes observed in the pyloric caeca or mid intestine. In the distal intestine, one of nine sampled fish fed nGM-soy showed moderate changes, two of nine sampled fish fed GM-soy showed changes, one with moderate and one with severe changes, and two of nine fish fed nGM-maize D2 had moderate changes. Using a monoclonal antibody against proliferating cell nuclear antigen (PCNA), cell proliferative responses to the experimental diets were assessed. In fish fed both soy diets, a significantly higher (P < 0.05) cell proliferation response was observed in the distal intestine concomitant with an increased localization of PCNA positive cells along the whole distal intestinal folds. The PCNA response among the nGM-soy group was significantly higher compared with all the other diet groups. In contrast, for fish exposed to dietary maize (type D) compared with fish fed the standard fishmeal, the soy-diets (GM-soy and nGM-soy) and maize (type P), a significantly lower (P < 0.05) cell proliferation response was observed in the distal intestine. Results indicated that the GM plant products investigated in this study, at about 12% inclusion level, were as safe as commercially available non-GM products, at least in terms of their effect on indices and histological parameters of the Atlantic salmon intestinal tract.

Photoperiod regulation of plasma growth hormone levels during induced smoltification of underyearling Atlantic salmon.

Earlier studies have established that increased daylength increases plasma growth hormone (GH) levels during spring smoltification of yearling Atlantic salmon. Recently, the Atlantic salmon aquaculture industry has started the production of underyearling ("summer") smolts. This involves fast juvenile growth on continuous light (24L), the transfer of juveniles over 8 cm in length to short day (12L) for 6 weeks in the summer, followed by transfer to 24L for another 6 weeks before transfer to seawater in late October. Three groups were studied in fresh water from July to the following May in order to elucidate the GH response to this photoperiod manipulation: one group was kept on 24L throughout (long-day group), while the other two groups were exposed to short day from July 15th. Of these, one was brought back onto long day on September 1st (winter group) while the other was kept on short day (short-day group). Plasma GH levels of the long-day group were around 1.6 ng/ml throughout the study. The short-day transfer suppressed GH levels to 0.7 ng/ml within 2 weeks (short-day and winter groups). The long-day transfer (winter group) increased GH levels to 11 ng/ml within 3 weeks, and this elevation of GH levels was sustained for about 3 months, before declining to pretreatment levels. The study demonstrates that underyearling Atlantic salmon react to increased daylength in a way similar to traditional yearling smolts. It also demonstrates for the first time that decreased daylength can suppress plasma GH levels in fish. It is concluded that winter photoperiod manipulation causes an out-of-season initiation and completion of the parr-smolt transformation of underyearling Atlantic salmon and that growth hormone plays a major role in this process.

beta-Hydroxybutyrate in developing nauplii of brine shrimp (Artemia franciscana K.) under feeding and non-feeding conditions.

Body content of beta-hydroxybutyrate, and individual dry mass, carbon content, and survival rate, were studied in developing nauplii of the brine shrimp Artemia franciscana K. from hatching to 96-97 h post hatching at 27 +/- 1 degrees C. The effect of two diets was studied in the experiment: Super Selco (SS) with a high lipid content; and Protein Selco (PS) with a high protein content. A starving group (S) was used as reference. The level of beta-hydroxybutyrate at hatching was 0.6 nmol.ind-1; it increased to 1.0-1.5 nmol.ind-1 in the SS- and S-groups, while in the PS-group it remained stable between 0.6-0.8 nmol.ind-1. At 60-80 h post hatch in the SS- and S-groups, the levels of beta-hydroxybutyrate were similar to the initial levels. The survival rate remained higher than 95% until 24 h post hatching in all groups. At the end of the experiment, the survival rate was 63% in the PS-group, 13% in the S-group and 3% in the SS-group. The Artemia nauplii individual dry mass and carbon content remained relatively stable in the SS-group; both parameters showed a significant increase in the PS-group and a significant decrease in the S-group. The results suggest that Artemia nauplii utilise ketone bodies as a fuel during development and growth, but that ketone catabolism may be overloaded by excessive lipid feeding resulting in increased mortality and possibly a ketotic acidosis.

Dietary carbohydrate utilization in cod (Gadus morhua): metabolic responses to feeding and fasting.

Moist diets with increasing amounts of carbohydrate (0.5%, 10% and 21% on a dry weight basis) were each fed to duplicate groups of cod (initial weight 370 g) for 8 weeks, after which all groups were fasted for 4 weeks. Protein energy was high and accounted for more than 70% of the gross energy content in all feeds, and the diets were maintained isocaloric by substituting lipid energy for carbohydrate energy. No indigestible binder was added. Excellent growth and feed conversion were obtained in all groups. After 4 weeks of fasting, fish previously fed diets with either 10% or 21% carbohydrate showed significantly higher weight loss than fish fed the diet without carbohydrate. Liver glycogen reached 10% of liver wet weight in fish fed diets containing 10% or 21% carbohydrate and 5% in fish receiving 0.5% carbohydrate after 8 weeks. Following 4 weeks of fasting, liver glycogen was reduced to similar levels in all fish. Plasma glucose levels 4 h after feeding were higher in fish fed the diets with 10% or 21% carbohydrate and plasma free amino acid levels (FAA) were lower, than in fish fed the diet containing 0.5% carbohydrate. Blood lactate concentrations were unaffected during the first 24 h after feeding. After 4 weeks of food deprivation, the levels were significantly reduced only in the 21% carbohydrate group. A link between glucagon and protein metabolism is suggested because plasma glucagon concentration followed the same pattern as the concentrations of plasma FAA throughout the study. Insulin and glucagon-like peptide (GLP) showed a covariation throughout the experiment. Reduced plasma insulin levels were seen after fasting concomitant with reduction in the levels of FAA and glucose. It is suggested that insulin secretion in cod is affected both by plasma FAA and glucose and that cod meets food deprivation by slowing down metabolism.

Insulin and glucagon family peptides in relation to activities of hepatic hexokinase and other enzymes in fed and starved Atlantic salmon (Salmo salar) and cod (Gadus morhua).

1. Plasma levels of insulin, glucagon, and glucagon-like peptide (Glp) were all reduced by starvation of salmon and cod. In the salmon the drop in Glp was larger than in insulin and glucagon. 2. After starvation the activity of hexokinase (EC 2.7.1.1) was increased in salmon liver, but decreased in cod liver. The salmon hepatic hexokinase activity was inversely correlated with the Glp/insulin ratio. 3. Activities of hepatic glycogen phosphorylase (EC 2.4.1.1) and phosphofructokinase (EC 2.7.1.11) were increased in starved as compared to fed salmon. In cod, starvation resulted in decreased or unchanged activity of phosphorylase. This discrepancy may be related to different degrees of environmental and handling stress. 4. Intraperitoneal injection of human insulin in salmon gave increased hepatic phosphorylase and hexokinase activities and reduced plasma levels of glucagon, Glp and endogenous fish insulin at sampling after 30 hr. 5. No differences in hepatic hexokinase activities or plasma hormone levels were observed between cod fed low and high carbohydrate diets. Apparently, regulation of glucose phosphorylation by dietary carbohydrate does not occur.