Digestive enzymes and metabolic profile of Pseudoplatystoma corruscans (Teleostei: Siluriformes) in response to diet composition

Digestive enzyme responsiveness to feeding and associated adjustments of metabolism can be used to derive nutritionally effective diet formulations. Juvenile pintado (Pseudoplatystoma corruscans) were fed different diets. After feeding, fish were killed and blood, liver and white muscle were collected to evaluate metabolites. Stomach along with anterior, middle and posterior intestine were sampled for enzyme analysis. Non-specific protease, trypsin, chymotrypsin, amylase and lipase were assayed. Crude protein (CP) did not induce proteolytic activity; highest protease activities were observed in the stomach. Amylase was higher in the stomach in fish feeding on diets containing 13-25% starch. Lipase activity was observed along the gastrointestinal tract, with the highest activities observed in the middle section. The metabolic profile of white muscle was not affected by CP. In contrast, some plasma and liver metabolites were altered concomitant with changes in the digestive enzymes. Amino acid catabolism was increased. Digestion in pintado was responsive to cornstarch, reflected in intermediary metabolism; proteolytic activities of the digestive tract seem to be sufficient to deal with large amounts of dietary protein. As a result, we are able to recommend a balance between protein and energetic compounds, such as lipids and carbohydrates, in the diet to optimize fish growth.     

Growth depression in socially subordinate rainbow trout Oncorhynchus mykiss: more than a fasting effect

To assess the effects of subordinate social status on digestive function, metabolism, and enzyme activity in salmonid fish, juvenile rainbow trout Oncorhynchus mykiss were paired with size-matched conspecifics (<1.5% difference in fork length) for 5 d. Fish that were fasted for 5 d and fish sampled directly from the holding tank were used as control groups. Both subordinate and fasted fish experienced significant decreases in intestine mass (P = 0.043), and the gall bladder showed marked and significant changes in both size (P = 0.004) and appearance. These findings suggest that the negative effect of social subordination on digestive function reflects in large part a lack of feeding. Hepatic phosphoenolpyruvate carboxykinase activity was significantly higher in subordinate fish relative to dominants, whereas subordinate hepatic pyruvate kinase activity was significantly lower; activities of both enzymes were significantly correlated with plasma cortisol concentrations and behavior scores. Dominant-subordinate differences in the activities of these enzymes were eliminated by administration of the glucocorticoid receptor antagonist RU486, underlining a role for circulating cortisol in eliciting the differences. Significant increases relative to control fish were also detected in red and white muscles from subordinate fish in the activities of protein catabolic enzymes (aspartate aminotransferase, alanine aminotransferase, glutamate dehydrogenase). These differences occurred in the absence of any change in plasma free amino acid or ammonia concentrations, supporting an enhanced turnover of amino acids in muscle in subordinate fish. The results support the hypothesis that changes in metabolism, beyond those elicited by low food consumption, may be responsible at least in part for the low growth rates typical of subordinate fish and that these changes may be related specifically to circulating cortisol levels in subordinate fish.     

The feeding response of diploid and triploid Atlantic salmon and brook trout

Pair and group experiments were conducted to determine whether differences exist in feeding success between juvenile diploid and triploid salmonids in a competitive situation. In the pair experiments, 22 pairs (one diploid and one triploid) of size-matched Quebec-strain brook trout Salvelinus fontinalis (7·2–46·3 g) were fed an unlimited number of pellets three times a day for 5 days. Dominance was assigned to the fish which ate the most pellets within each pair. In the group experiments, groups of three diploid and three triploid size-matched fish were fed a restricted ration three times a day for 5 days. Hierarchical rank within the group was assigned based on the number of pellets consumed by each fish. The group experiment was repeated 10 times with Atlantic salmon Salmo salar (5·1–62·7 g), Quebec-strain brook trout (11·8-110·8 g), and large UNB-strain brook trout (18·2–33·0 g), and 12 times with smaller UNB-strain brook trout (0·6–2·0 g). A statistically significant difference in rank between ploidies was found only for the smaller UNB-strain brook trout in the group experiments, with diploids dominant over triploids. This suggests that there may be a difference in competitive feeding success between diploid and triploid brook trout early in development, but that this difference diminishes as the fish grow.     

Effect of temperature on heart rate in diploid and triploid brook charr, Salvelinus fontinalis, embryos and larvae

Increased cell size in triploid fish likely affects rates of respiratory gas exchange. Respiratory deficiencies can be addressed in fish by adjustments in cardiac output, through changes in heart rate and stroke volume. The aim of this study was to determine whether heart rate differs between triploid and control (diploid) brook charr, Salvelinus fontinalis, at embryo-larval stages, when the heart is easily visible and the fish are relatively inactive. Heart rate was measured at 6, 9 and 12 degrees C at three developmental stages: eyed-egg, hatch and yolk absorption. Heart rate was unaffected by ploidy, but increased with temperature and age from a low of 43.4+/-2.2 beats/min (6 degrees C, eyed egg) to a high of 73.3+/-1.5 beats/min (12 degrees C, yolk absorption). The Q(10) for heart rate was unaffected by ploidy and age, but decreased with temperature from 1.99+/-0.28 at 6-9 degrees C to 1.72+/-0.17 at 9-12 degrees C. Triploid brook charr thus do not use adjustments in heart rate as a mechanism to deal with the physiological consequences of altered haematology at embryo-larval stages.     

Metabolic adaptation of Escherichia coli during temperature-induced recombinant protein production: 1. Readjustment of metabolic enzyme synthesis

The metabolic burden and the stress load resulting from temperature-induced production of human basic fibroblast growth factor is connected to an increase in the respiratory activity of recombinant Escherichia coli, thereby reducing the biomass yield. To study the underlying changes in metabolic enzyme synthesis rates, the radiolabeled proteom was subjected to two-dimen- sional gel electrophoresis. After temperature-induction, the cAMP-CRP controlled dehydrogenases of the pyruvate dehydrogenase complex and the tricarboxylic acid cycle (LpdA and SdhA) were induced four times, reaching a maximum 1 h after the temperature upshift. The more abundant tricarboxylic acid cycle dehydrogenases (Icd and Mdh) were initially produced at reduced rates but regained preshift rates within 30 min. The adenylate energy charge dropped immediately after the temperature upshift but recovered within 1 h. Similar profiles in dehydrogenase synthesis rates and adenylate energy charge were found in a control cultivation of a strain carrying the "empty" parental expression vector. Although both strains exhibited significant differences in growth pattern and respiration rates after the temperature upshift, the adaptation of the energetic state of the cells and the synthesis of enzymes from the energy-generating catabolic pathway did not seem to be affected by the strong overproduction of the recombinant growth factor. In contrast, the synthesis rates of enzymes belonging to the biosynthetic machinery, e.g., translational elongation factors, decreased more strongly in the culture synthesizing the recombinant protein. In control and producing culture, synthesis rates of elongation factors paralleled the respective growth rate profiles. Thus, cells seem to readjust their metabolic activities according to their energetic requirements and, if necessary, at the cost of their biosynthetic capabilities.     

Morphological comparisons of diploid and triploid hybrid grass carp, Ctenopharyngodon idella♀×Hypophthalmichthys nobilis♂

Morphological and karyological comparisons of 25–28 hybrid grass carp from each of 3 year-classes (1979–1981) indicated that diploid fish resulted during 1979 (82.1%) and 1980 (76.0%) the others being triploid. All of the fish tested from the 1981 year-class were triploid. Most triploid fish differed from diploid fish by having a faster growth rate, fewer scales in the lateral line and transverse series below the lateral line, a relatively longer gut, and fewer deformities.     

Modulation of digestive enzyme activities during ontogeny of Labeo rohita larvae fed ascorbic acid enriched zooplankton

The effect of supplementation of ascorbic acid through enriched zooplankton [10%, 20% and 30% ascorbyl palmitate (AP) inclusion in diet of zooplankton] on different digestive enzyme activities during ontogeny of Labeo rohita larvae was studied from 4 day to 15 day post hatch. Ascorbic acid (AA) content in different groups of unenriched (8.6+/-0.71) and enriched zooplankton were, 750+/-29.3, 1409.1+/-45.5, 2009.21+/-199.2 mug/g respectively on dry matter basis with differences (P<0.05) between the treatments. A difference (P<0.05) was found in tissue AA level in different dietary groups. Low amylase, protease, lipase and alkaline phosphatase activities were present in rohu larvae from the mouth opening stage which showed increasing trend with the age of larvae and increasing dietary AA content. A clear dose-dependent modulation of digestive enzyme activities in response to 10%, 20% and 30% AP enriched zooplankton feeding was evidenced from positive correlations between dietary AA content with magnitude of elevation of enzyme activity in different groups. There were 57, 55, 29.2 and 2 fold increases in amylase activity; 7.35, 7.02, 4.43 and 2.73 fold increases in protease activity; 45.636, 41.50, 19.83 and 13.69 fold increases in lipase activity and 6, 5, 3, and 2 fold increases in alkaline phosphatase activity observed in the 15th day post hatch larvae fed 20%, 30%, 10%AP enriched and normal zooplankton respectively, than 4-day post hatch larvae of the respective groups. Enzyme activities were also positively correlated with specific growth rates of wet weight of rohu larvae at the 15th day post hatch. Increased AA might have played an important role in advancing morphological transformation of the digestive tract, protecting gastric mucosa and accelerating growth by the process of tissue formation, which necessitated the requirement of more nutrient thereby, increasing digestive enzyme activity. The regulatory role of AA in the modulation of different digestive enzymes activity and its physiological consequences of nutrient digestibility and utilization during ontogenesis could be extrapolated for better nutrient management of the larvae.     

Growth and nutrient utilization by 0 + and 1 + triploid rainbow trout, Oncorhynchus my kiss

Growth performance, protein and energy utilization in triploid rainbow trout juveniles of two yearclasses, 0 + and 1 +, were compared to diploid control groups. Growth rate, food conversion and protein efficiency ratios of triploid rainbow trout did not differ (0 + juveniles) or were slightly lower (1 + juveniles) than the diploid control. Proximate body composition was also not affected by ploidy.
Apparent digestibility coefficients (ADC) ofprotein and ofenergy, oxygen uptake, and ammonia excretion rates were not different between the two groups. Nitrogen and energy balance studies showed that digestive and metabolic utilization of the diets did not differ in triploid and diploid rainbow trout juveniles.     

Hematologic and enzymatic analysis of Ctenopharyngodon idella x Hypophthalmichthys nobilis F1 hybrids

1. Erythrocyte counts, hemoglobin concentrations and hematocrit values were determined for diploid and triploid Ctenopharyngodon idella X Hypophthalmichthys nobilis hybrids and the parental species. 2. Comparisons of diploid and triploid hybrids with the parental species revealed low erythrocyte counts for triploids, high mean corpuscular hemoglobin values for triploids, elevated hematocrits for diploids and triploids and similar hemoglobin concentrations for all fish. 3. Alkaline phosphatase, aldolase, and lactate dehydrogenase specific activities were determined spectrophotometrically. Levels of specific activity of these enzymes in the hybrids were consistently elevated above that of the parental species. These higher levels of enzyme activities in hybrids were probably the result of a breakdown in gene regulation.     

Mucous cell types in the branchial epithelium of the euryhaline fish Poecilia vivipara

On transfer to sea water for 45 days, the return of appetite was later and growth rates tended to be lower for triploid Atlantic salmon, Salmo salar , reared together with diploid Atlantic salmon. All mortalities comprised of triploid salmon (29%) and were attributable to failed smolt syndrome. No correlation was found between the growth of diploid or triploid fish in fresh water and their subsequent growth on transfer to sea water.     

The kinetics of the hypoferraemic response and changes in levels of alternative complement activity in diploid and triploid Atlantic salmon, following injection of lipopolysaccharide.

To study any possible effects of triploidy on the kinetics of the response of two non-specific disease factors, full sibling diploid and triploid Atlantic salmon were injected intraperitoneally with either lipopolysaccharide (1 mg kg(-1) body weight) or saline. Individually marked fish were repetitively blood sampled for up to 19 days. Total serum protein concentrations remained constant throughout the experiment indicating that the sampling regime did not cause haemodilution. The alternative complement pathway activity (measured by the titre of haemolytic activity against rabbit erythrocytes) in the serum of saline injected fish remained constant but in LPS-injected fish it fell to barely detectable levels 2 days after injection, but recovered to pre-treatment levels by about day 5. Triploid fish took slightly longer to reach full recovery levels than diploids. All groups of fish showed a hypoferraemic response, suggesting that the sampling regime was at least partially responsible. However, the response was more rapid and pronounced in the LPS-injected fish. In the latter, serum iron concentrations decreased to very low levels by day 2 post-injection in the diploid fish and by day 3 in the triploid fish. Pre-treatment iron levels were re-established by about 15 days post-injection in all groups. The data show only slight differences between the diploid and triploid fish, but the longer time taken for the triploids to recover complement activity and the slower onset of the hypoferraemic response following injection of LPS, suggest that they may be at a disadvantage compared with their diploid siblings in their defence against bacterial infections.     

Erythrocyte enzyme activities in diploid and triploid salamanders (Pleurodeles waltlii) of both sexes

Studies of several enzyme activities and of some metabolites in erythrocytes of the newt Pleurodels waltilii are described. The results indicate that differences exist between enzyme activities in males and females and in diploid and triploid animals. Females have higher erythrocyte enzyme activities than males. However, triploid animals have lower enzyme activites in their erythrocytes than diploid animals; this observation is more striking in females than in males.     

The activity of several components of the innate immune system in diploid and triploid turbot

The use of triploid fish may be of interest in research, e.g. study of how this condition affects the size and activity of cells. In addition, triploid fish are sterile and production of triploids in fish species that are marketed after reaching sexual maturity may be of economic interest. In the present study, the effects of triploidy on the activity of several components of the innate immune system of turbot (Psetta maxima L) were determined. Triploid turbot had bigger cells (erythrocytes and neutrophils) but the number of blood erythrocytes, leucocytes and thrombocytes was lower than in diploid fish. The differential cell count was similar in both types of fish. The respiratory burst and the phagocytic activities were higher in neutrophils of triploid turbot. However, because the number of neutrophils was higher in diploids, the total respiratory burst activity and the phagocytosis per microliter of blood was similar in both types of fish. No differences were found in serum complement, lysozyme or bactericidal activities. The results indicate that the activities of the humoral components of the innate immune system tested are similar in diploid and triploid fish and that the lower leucocyte number found in triploids is compensated for by higher cell activity.     

Comparative growth performance of diploid and triploid European sea bass over the first four spawning seasons

During their 3–4 first years of life, triploid sea bass Dicentrarchus labrax grew in a similar fashion to diploids in fork length but more slowly than diploids ( P <0·05) in body weight, even when the diploids reached full sexual maturity. However, from 48–53 months of age triploids exhibited non-significantly higher instantaneous growth rates, and thus when fish were 4 years or older, differences in weight with diploids were no longer apparent, suggesting that triploidy could be of benefit in the culture of large (>1 kg) sea bass. The condition factor was reduced in both ploidies during the spawning season which took place in winter when the temperature was low. These observations suggest that any growth advantage in triploids, which were functionally sterile, may be offset by unfavourable environmental conditions. Thus, the potential gain of triploid fish, because they do not direct energy to gonadal growth, could not overcome the effects of low temperature on somatic growth, which coincided with the spawning season. This suggests that the low growth of this species during winter is more a consequence of low temperature than of the energetic cost associated with reproduction. On the other hand, the lower hepatosomatic index in triploid females in contrast to diploid females might be indicative of the lack of gonadal oestradiol-mediated hepatic synthesis of vitellogenin. Also, erythrocyte and haematocrit measurements showed an increased nuclear and cellular volume in triploids, but with similar cell numbers to those of diploids, respectively.     

MicroRNA Alternations in the Testes Related to the Sterility of Triploid Fish

The knowledge of understanding the molecular traits of the sterile triploid fish is sparse. Herein, we analyzed the microRNA (miRNA) alternations in the testes of the sterile triploid fish produced by crossing the tetraploid fish with the diploid fish, compared with those of tetraploids and diploids used as the controls. A total of 136, 134, and 142 conserved miRNAs and 105, 112, and 119 novel miRNAs were identified in the diploid, triploid, and tetraploid fish, respectively. The genes targeted by the differentially expressed miRNAs were identified and were enriched in the GO term cell surface receptor signaling pathway, cellular process, G-protein coupled receptor signaling pathway, and metabolic process. KEGG pathway enrichment was also assessed to evaluate the target genes with differentially expressed miRNAs and these genes were enriched in four pathways (synthesis and degradation of ketone bodies, pentose and glucuronate interconversions, cyanoamino acid metabolic process, and ascorbate and aldarate metabolism). Nine differentially expressed miRNAs were verified by quantitative real-time PCR analysis (qPCR). The upregulated miRNAs in triploids, including miR-101a, miR-199-5p, miR-214, miR-222, and miR-193a, showed the same results with high-throughput sequencing. Among the selected downregulated miRNAs, miR-7b and miR-153b had significantly lower expression levels in triploids. Dnah3 and Tekt1 genes targeted by miR-199-5p showed lower expression in triploids by qPCR. These verified differentially expressed miRNAs may participate in testicular development and sperm activity by targeting functional genes, which were identified with differential expression in the triploid. This evidence provides insights into the epigenetic regulatory mechanisms of sterility in triploid cyprinids.     

Triploidy in rainbow trout determined by computer-assisted analysis

This study was designed to assess the use of a computer-assisted system based on erythrocyte measurements as a possible alternative to flow cytometry for identifying triploid rainbow trout (Oncorhynchus mykiss). Blood smears were prepared from 26 triploid and 26 diploid specimens, as determined by flow cytometry after staining blood cells with propidium iodide. The cell and nucleus lengths of 10 erythrocytes were determined in each fish. This was followed by discriminatory analysis to distinguish between diploids and triploids based on their score profiles. Triploid trout showed significantly larger erythrocyte cell and nucleus measurements than their diploid counterparts (N=52; P<0.0001). Erythrocyte length correctly identified 100% of the fish specimens as diploid or triploid, while nucleus length was a less accurate predictor of the level of ploidy. Our findings validate the potential use of computer-assisted analysis for this purpose.     

Biochemical and morphological correlates of growth in diploid and triploid rainbow trout

Diploid and triploid rainbow trout contained the same amount of RNA per unit weight of white muscle tissue at a given growth rate. This finding, along with a previous finding that the protein content of triploid muscle tissues is not different from that of the diploids, indicates that triploid nuclei produce more RNA than their diploid counterparts. This increased production of RNA is probably due to the expression of the extra set of chromosomes in triploid nuclei. RNA-based growth correlates and liver somatic index ( I _H) were valid predictors of growth for both triploid and diploid fish. RNA concentration appeared to be better than RNA/DNA and RNA/protein in predicting growth.     

Effect of normal and waxy maize starch on growth, food utilization and hepatic glucose metabolism in European sea bass (Dicentrarchus labrax) juveniles

We determined the effect of dietary starch on growth performance and feed utilization in European sea bass juveniles. Data on the dietary regulation of key hepatic enzymes of the glycolytic, gluconeogenic, lipogenic and amino acid metabolic pathways (hexokinase, HK; glucokinase, GK; pyruvate kinase, PK; fructose-1,6-bisphosphatase, FBPase; glucose-6-phosphatase, G6Pase; glucose-6-phosphate dehydrogenase, G6PD; alanine aminotransferase, ALAT; aspartate aminotransferase, ASAT and glutamate dehydrogenase, GDH) were also measured. Five isonitrogenous (48% crude protein) and isolipidic (14% crude lipids) diets were formulated to contain 10% normal starch (diet NS10), 10% waxy starch (diet WS10), 20% normal starch (diet NS20), 20% waxy starch (diet WS20) or no starch (control diet). Another diet was formulated with no carbohydrate, and contained 68% crude protein and 14% crude lipids (diet HP). Each experimental diet was fed to triplicate groups of 30 fish (initial weight: 23.3 g) on an equivalent feeding scheme for 12 weeks. The best growth performance and feed efficiency were achieved with fish fed the HP diet. Neither the level nor the nature of starch had measurable effects on growth performance of sea bass juveniles. Digestibility of starch was higher with waxy starch and decreased with increasing levels of starch in the diet. Whole-body composition and plasma metabolites, mainly glycemia, were not affected by the level and nature of the dietary starch. Data on enzyme activities suggest that dietary carbohydrates significantly improve protein utilization associated with increased glycolytic enzyme activities (GK and PK), as well as decreased gluconeogenic (FBPase) and amino acid catabolic (GDH) enzyme activities. The nature of dietary carbohydrates tested had little influence on performance criteria.