Allozyme diversity in Australian rainbow trout, Oncorhynchus mykiss

Rainbow trout, Oncorhynchus mykiss (Walbaum), were first introduced into Australia over 100 years ago, and forms the basis of important recreational inland fisheries and an aquaculture industry in south-eastern Australia. This paper investigates the genetic variation within and between samples of Australian rainbow trout using allozyme electrophoresis. The levels of genetic diversity within Australia do not show marked differences from those observed in hatchery and wild populations from throughout North America, New Zealand and South Africa, but there is evidence for the loss of some rare alleles during translocation from California to Australia via New Zealand. No appreciable difference in genetic diversity was apparent between hatchery and self-sustaining wild populations of rainbow trout from mainland Australia. However, significant differences in allelic frequencies were observed, with consistent genetic differences between Victorian and New South Wales samples most likely reflecting state-based hatchery and stocking policies.

Determination of astaxanthin steroisomers and colour attributes in flesh of rainbow trout (Oncorhynchus mykiss) as a tool to distinguish the dietary pigmentation source

The presence of carotenoids in animal tissue reflects their sources along the food chain. Astaxanthin, the main carotenoid used for salmonid pigmentation, is usually included in the feed as a synthetic product. However, other dietary sources of astaxanthin such as shrimp or krill wastes, algae meal or yeasts are also available on the market. Astaxanthin possesses two identical asymmetric atoms at C-3 and C-3' making possible three optical isomers with all-trans configuration of the chain: 3S,3'S, 3R,3'S, and 3R,3'R. The distribution of the isomers in natural astaxanthin differs from that of the synthetic product. This latter is a racemic mixture, with a typical ratio of 1:2:1 (3S,3'S:3R,3'S:3R,3'R), while astaxanthin from natural sources has a variable distribution of the isomers deriving from the different biological organism that synthesized it. The high-performance liquid chromatographic (HPLC) analysis of all-trans isomers of astaxanthin was performed in different pigment sources, such as red yeast Phaffia rhodozyma, alga meal Haematococcus pluvialis, krill meal and oil, and shrimp meal. With the aim to investigate astaxanthin isomer ratios in flesh of fish fed different carotenoid sources, three groups of rainbow trout were fed for 60 days diets containing astaxanthin from synthetic source, H. pluvialis algae meal and P. rhodozyma red yeast. Moreover, the distribution of optical isomers of astaxanthin in trout purchased on the Italian market was investigated. A characteristic distribution of astaxanthin stereoisomers was detected for each pigment sources and such distribution was reproduced in the flesh of trout fed with that source. Colour values measured in different sites of fillet of rainbow trout fed with different pigment sources showed no significant differences. Similarly, different sources of pigment (natural or synthetic) produced colour values of fresh fillet with no relevant or significant differences. The coefficient of distance computed amongst the feed ingredient and the trout fillet astaxanthin stereoisomers was a useful tool to identify the origin of the pigment used on farm.

Expression of natriuretic peptide receptor mRNA and functional response to atrial natriuretic peptide and C-type natriuretic peptide in rainbow trout (Oncorhynchus mykiss) head kidney leucocytes

The stimulatory effect of vasomodulatory natriuretic peptide hormones on macrophages and peripheral blood leucocytes in mammals is well-established. However, the relationship in lower vertebrates has not been characterised. Expression of atrial natriuretic peptide, ventricular natriuretic peptide and C-type natriuretic peptide-1, and the guanylyl cyclase-linked (GC) natriuretic peptide receptor-A and -B-type receptors (NPR-A and NPR-B, respectively) was determined by PCR from the mRNA of rainbow trout head kidney leucocytes yielding gene fragments with 100% homology to the same respective natriuretic peptide and NPR-A and -B sequences obtained from other rainbow trout tissues. A mixed population of isolated rainbow trout head kidney leucocytes was stimulated in vitro with trout atrial natriuretic peptide (specific NPR-A agonist) and trout C-type natriuretic peptide (NPR-A and -B agonist) as well as the cGMP agonist 8-bromo-cGMP or the GC inhibitor 8-bromo-phenyl-eutheno-cGMP. Respiratory burst was stimulated by trout atrial natriuretic peptide, trout C-type natriuretic peptide-1 and 8-bromo-cGMP in a dose dependant manner with the highest activity as a result of stimulation with trout C-type natriuretic peptide-1 in excess of that achieved by phorbol myristate acetate (PMA). Equimolar concentrations of the inhibitor, inhibited the respiratory burst caused by the natriuretic peptides and 8-bromo-cGMP. The natriuretic peptide receptors on rainbow trout head kidney leucocytes appear to have a stimulatory function with regard to respiratory burst that is activated through a cGMP second messenger pathway and the natriuretic peptides expressed in the head kidney leucocytes may well act in a paracrine/autocrine manner.

Over-winter lipid depletion and mortality of age-0 rainbow trout (Oncorhynchus mykiss)

In this study we identify the size-dependent risk of winter starvation mortality as a strong selective pressure on age-0 rainbow trout (Oncorhynchus mykiss) that could promote the risk-taking behaviour and allocation of energy to lipids previously observed in young trout cohorts. Age-0 trout subjected to simulated winter starvation conditions gradually depleted lipid reserves to a critical minimum lipid content below which death occurred. Small fish with lower lipid content exhausted lipid reserves earlier, and experienced high mortality rates sooner, than larger fish with greater lipid content. Consequently, winter starvation endurance was dependent upon size-dependent lipid reserves and winter duration. To validate the laboratory findings in the field, we stocked several size classes of hatchery-raised trout with known lipid content at the start of winter into two experimental lakes, and estimated survival and lipid depletion at winter's end. Larger age-0 trout had greater initial lipid reserves than smaller trout. Individuals depleted most of their lipid reserves over the winter, and experienced mortality that ranged from just under 60% for the largest individuals to just over 90% of the smallest individuals. Many survivors had lipid contents near, but none were below, the minimum lipid content determined in the laboratory.

Stress response of juvenile rainbow trout (Oncorhynchus mykiss)to chemical cues released from stressed conspecifics

The objective of this study was to determine whether exposure of rainbow trout (Oncorhynchus mykiss) to water containing a stressed trout or skin extract from stressed and non-stressed trout would elicit a stress response in conspecifics. Juvenile rainbow trout were exposed for 1 hour to water containing a stressed fish, homogenized skin extracts from a non-stressed fish, skin extract from a stressed fish and water with none of these factors. The stress response was measured over a 24-h period (1, 6, 12, 24 h after exposure). Plasma cortisol levels increased at 12 h in fish exposed to water from a stressed fish and skin extract from a stressed fish. Plasma glucose and hepatic hsp70 levels were not affected by treatments. The results suggest that rainbow trout elicit a stress response when exposed to stress-related alarm cues released from conspecifics.

The orexinergic system in rainbow trout Oncorhynchus mykiss and its regulation by dietary lipids

In this study, we report the distribution of orexin A (OXA), orexin B (OXB), and orexin receptor (OX2R) immunoreactive (ir) cells in the hypothalamus and gastrointestinal tract of Oncorhynchus mykiss fed diets with different dietary fatty acid compositions. Trout were fed five iso-energetic experimental diets containing fish oil, or one of four different vegetable oils (olive, sunflower, linseed, and palm oils) as the added dietary lipid source for 12 weeks. OXA, OXB, and OX2R immunoreactive neurons and nervous fibers were identified in the lateral and ventro-medial hypothalamus. OXA, OXB, and OX2R ir cells were found in the mucosa and glands of the stomach and in the mucosa of both the pyloric cecae and intestine. OX2R ir cells were localized in the mucosa layer of both the pyloric cecae and intestine. These immunohistochemical (IHC) results were confirmed via Western blotting. Antibodies against preproorexin (PPO) crossreacted with a band of ∼16 kDa in the hypothalamus, stomach, pyloric cecae, and intestine. Antibodies against OX2R crossreacted with a band of ∼38 kDa in the hypothalamus, pyloric cecae, and intestine. The presence and distribution of OXA, OXB, and OX2R ir cells in the hypothalamus and gastrointestinal tract did not appear to be affected by dietary oils. The presence of orexin system immunoreactive cells in the stomach, pyloric cecae, and intestine of rainbow trout, but not in the enteric nervous system, could suggest a possible role of these peptides as signaling of gastric emptying or endocrine modulation, implying a main local action played by orexins.

Isolation and functional Characterisation of a fads2 in Rainbow Trout (Oncorhynchus mykiss) with Δ5 desaturase activity

Rainbow trout, Oncorhynchus mykiss, are intensively cultured globally. Understanding their requirement for long-chain polyunsaturated fatty acids (LC-PUFA) and the biochemistry of the enzymes and biosynthetic pathways required for fatty acid synthesis is important and highly relevant in current aquaculture. Most gnathostome vertebrates have two fatty acid desaturase (fads) genes with known functions in LC-PUFA biosynthesis and termed fads1 and fads2. However, teleost fish have exclusively fads2 genes. In rainbow trout, a fads2 cDNA had been previously cloned and found to encode an enzyme with Δ6 desaturase activity. In the present study, a second fads2 cDNA was cloned from the liver of rainbow trout and termed fads2b. The full-length mRNA contained 1578 nucleotides with an open reading frame of 1365 nucleotides that encoded a 454 amino acid protein with a predicted molecular weight of 52.48 kDa. The predicted Fads2b protein had the characteristic traits of the microsomal Fads family, including an N-terminal cytochrome b5 domain containing the heme-binding motif (HPPG), histidine boxes (HDXGH, HFQHH and QIEHH) and three transmembrane regions. The fads2b was expressed predominantly in the brain, liver, intestine and pyloric caeca. Expression of the fasd2b in yeast generated a protein that was found to specifically convert eicosatetraenoic acid (20:4n-3) to eicosapentaenoic acid (20:5n-3), and therefore functioned as a Δ5 desaturase. Therefore, rainbow trout have two fads2 genes that encode proteins with Δ5 and Δ6 desaturase activities, respectively, which enable this species to perform all the desaturation steps required for the biosynthesis of LC-PUFA from C18 precursors.

Nutritional regulation of long-chain PUFA biosynthetic genes in rainbow trout (Oncorhynchus mykiss)

Most studies on dietary vegetable oil in rainbow trout (Oncorhynchus mykiss) have been conducted on a background of dietary EPA (20 : 5n-3) and DHA (22 : 6n-3) contained in the fishmeal used as a protein source in aquaculture feed. If dietary EPA and DHA repress their endogenous synthesis from α-linolenic acid (ALA, 18 : 3n-3), then the potential of ALA-containing vegetable oils to maintain tissue EPA and DHA has been underestimated. We examined the effect of individual dietary n-3 PUFA on the expression of the biosynthetic genes required for metabolism of ALA to DHA in rainbow trout. A total of 720 juvenile rainbow trout were allocated to twenty-four experimental tanks and assigned one of eight diets. The effect of dietary ALA, EPA or DHA, in isolation or in combination, on hepatic expression of fatty acyl desaturase (FADS)2a(Δ6), FADS2b(Δ5), elongation of very long-chain fatty acid (ELOVL)5 and ELOVL2 was examined after 3 weeks of dietary intervention. The effect of these diets on liver and muscle phospholipid PUFA composition was also examined. The expression levels of FADS2a(Δ6), ELOVL5 and ELOVL2 were highest when diets were high in ALA, with no added EPA or DHA. Under these conditions ALA was readily converted to tissue DHA. Dietary DHA had the largest and most consistent effect in down-regulating the gene expression of all four genes. The ELOVL5 expression was the least responsive of the four genes to dietary n-3 PUFA changes. These findings should be considered when optimising aquaculture feeds containing vegetable oils and/or fish oil or fishmeal to achieve maximum DHA synthesis.

Discrimination of origin of farmed trout by means of biometrical parameters, fillet chemical composition and flavour volatile compounds

To date it is well known that the quality of farmed trout is affected by diet composition, by feeding regime, by husbandry practices and by rearing conditions and environment. The trout processing industry and the large-scale retail trade, in consideration of the wide variability of trout quality and characteristics, have imposed, or will soon impose, quality criteria for the end product. Moreover, recent food scares and the malpractices of some food producers have increased public requests
for traceability. The aim of the present study was to evaluate the main chemical quality and the biometrical characteristics of rainbow trout produced in three different farms in Italy (two intensive farms, located one on mountain and one on plain, and an extensive farm in which fish fed only on naturally available nutrients) and to establish whether farmed trout
origins could be differentiated by these parameters. Trout farmed in the intensive mountain farm (IMF) showed the highest crude lipid content in the fillets and the fatty acids of their fillets were characterized by the highest percentage of MUFA. Trout farmed in the intensive plain farm (IPF) were characterized by low dressing percentage, and the lipid of their fillets
was rich in n-6 fatty acids. Trout stocked for the last year of their life in the extensive farm (EF) were leaner both in the carcass and in the fillets. The analysis of flavor volatile compounds showed some differences in the bouquet design, particularly differences in the amounts of n-3 and n-6 derivates volatile aldehydes and alcohols. All data significantly different
(P<0.05) were subjected to Linear Discriminant Analysis (LDA) and 8 variables were chosen to create two discriminant equations generating a strong prediction model for classification of farmed trout respective to their origins.

Effects of the extensive culture system as finishing production strategy on biometric and chemical parameters in rainbow trout

The effect of finishing extensive farming period, to reduce fat content and manipulate the fatty acid profile of fish muscle, was evaluated in rainbow trout. Fish were stocked in an artificial lake, in which fish were fed only on naturally available nutrients with no supply of artificial feed, for different lengths of time from 0 to 120 days. No weight loss was noted during the whole finishing period while total length increased from 228±7 to 269±3 mm and the condition factor decreased from 1.41±0.04 to 0.89±0.02. The total fat content of the fillets decreased considerably from 4.7±0.6% at the beginning to 2.4±0.4% and 0.7±0.2% after 45 and 120 days respectively. Fillet fatty acid composition was affected by the time of stocking in the extensive farm. In contrast to the reduction in C18:1n-9, C18:2n-6, total monounsaturated fatty acid and total n-6 percent values, an increase in the C20:5n-3, C22:6n-3, total polyunsaturated fatty acid and total n-3 percent values was observed. It was shown that while other finishing strategies for salmonids have some disadvantages, the extensive culture system seems to be a potentially useful tool for increasing the general quality of the end product.