A reappraisal of activity metabolism in Atlantic cod (Gadus morhua)

Atlantic cod ( Gadus morhua ) were forced to swim in a swim tunnel respirometer until fatigued; oxygen consumption rate (O₂) was measured during swimming at incremental speeds until the fish was exhausted and during recovery from exhaustion. Maximal oxygen consumption (O_2max) occurred during maximal activity as has been found for other fish species, but at odds with the current paradigm for Atlantic cod. Earlier experiments had drawn the conclusion that O_2max in Atlantic cod occurs during recovery from exhaustive exercise. We found no support for this paradigm in our experiments and we propose that the respiratory physiology of Atlantic cod is not unlike that of other fishes.     

Persistent growth effects of temperature and photoperiod in Atlantic cod Gadus morhua

Short-term environmental manipulations during the early juvenile stage have a large impact on harvesting size of Atlantic cod Gadus morhua nearly 3 years later. A group of juvenile Atlantic cod (initial mass 9·5 g) were reared for 3 months under simulated natural photoperiod or continuous light, and a range of temperatures (7, 10, 13 and 16° C, and a group called T-step, i.e. with temperature reduced successively from 16 to 13 and 10° C). After termination of the laboratory trial, the fish were moved to sea pens and reared at ambient conditions for 30 months before harvesting in June 2006. Observed growth gain from the 3 month laboratory trial was still persistent following the 30 months of sea-pen ongrowing. The T-step group displayed 15, 13, 1 and 10% superior mass gain respectively than the groups initially at 7, 10, 13 and 16° C at harvest in June 2006. Similarly, rearing under continuous light during the initial 3 month period during the early juvenile stage resulted in 1–9% larger size at harvesting compared to fish reared at simulated natural photoperiod. Gonado-somatic and hepato-somatic indices were similar in all groups. Contribution to the understanding of the mechanism behind size variation in adult fish can have wide range applications for Atlantic cod fisheries and aquaculture.     

Expression of the oligopeptide transporter, PepT1, in larval Atlantic cod (Gadus morhua)

The intestinal absorption of di- and tri-peptides generally occurs via the oligopeptide transporter, PepT1. This study evaluates the expression of PepT1 in larval Atlantic cod (Gadus morhua) during the three weeks following the onset of exogenous feeding. Larval Atlantic cod were fed either wild captured zooplankton or enriched rotifers. cDNA was prepared from whole cod larvae preceding first feeding and at 1000 each Tuesday and Thursday for the following three weeks. Spatial and temporal expression patterns of PepT1 mRNA were compared between fish consuming the two prey types using in situ hybridization and quantitative real-time PCR. Results indicated that PepT1 mRNA was expressed prior to the onset of exogenous feeding. In addition, PepT1 was expressed throughout the digestive system except the esophagus and sphincter regions. Expression slightly increased following first-feeding and continued to increase throughout the study for larvae feeding on both prey types. When comparing PepT1 expression in larvae larger than 0.15-mg dry mass with expression levels in larvae prior to feeding, no differences were detected for larvae fed rotifers, but the larvae fed zooplankton had significantly greater PepT1 expression at the larger size. In addition, PepT1 expression in the zooplankton fed larvae larger than 0.15-mg dry mass had significantly greater expression than rotifer fed larvae of a similar weight. Switching prey types did not affect PepT1 expression. These results indicate that Atlantic cod PepT1 expression was slightly different relative to dietary treatment during the three weeks following first-feeding. In addition, PepT1 may play an important role in the larval nutrition since it is widely expressed in the digestive tract.     

Minisatellite DNA fingerprints of Atlantic cod (Gadus morhua)

The human minisatellite probes 33.6 and 33.15 cross–hybridized to Hae III and Hinf I digested cod DNA, revealing complex fragment patterns in both Arctic and coastal cod. The DNA fingerprints were highly polymorphic, individual specific and stable in the germline. The potential applications of multi locus probes in cod research are discussed.     

Diclidophora merlangi (Monogenea: Diclidophoridae) on Atlantic cod, Gadus morhua

Diclidophora merlangi (Kuhn, in Nordmann, 1832), as species specific to whiting, Merlangius merlangus (Linnaeus, 1758), is reported and described for the first time on the gills of Atlantic cod Gadus morhua (Linnaeus, 1758). Of the about 1,200 cod examined in this article, only 2 specimens of D. merlangi occurred on 2 fish, suggesting that they represented accidental infections. The 2 specimens showed specific traits of D. merlangi, but their size was considerably smaller than that of D. merlangi on whiting. Principal-component analyses showed that the morphology of the specimens on cod was more similar to that of D. merlangi than those of other congeneric species from the North Atlantic, additionally supporting their assignment to D. merlangi. Quantitative data documenting the potential reproductive consequences of D. merlangi occurring on cod is also provided. The testes and germaria of the specimens on cod were noticeably smaller than those of D. merlangi on whiting, but the number of testes was similar and spermatozoa were observed in the sperm duct. By contrast, the aspect and smaller size of the oocytes suggested that D. merlangi on cod could not produce viable ova, although 1 specimen exhibited an egg with normal sized and shaped capsule.     

Substrate selection by juvenile Atlantic cod (Gadus morhua): effects of predation risk

Although predator avoidance has been proposed as one possible factor influencing the distribution of fish among substrate types, no study has addressed this question directly. Groups of juvenile Atlantic cod were offered a choice between pairs of the following three substrates: sand, gravel-pebble and cobble. Their distribution on these substrates was compared prior to, during and following exposure to a predator (i.e. a larger conspecific). With no apparent risk of predation, juvenile cod preferred sand or gravel-pebble. When cobble was present, juveniles hid in the interstitial spaces of this substrate in the presence of a predator. With no cobble present, juveniles showed no preference between sand and gravel-pebble, and did not seek refuge from predation in association with these substrates. Following exposure to a predator (i.e. 2.5 h later) larger juvenile cod again showed a preference for the finer-grained substrates, but smaller individuals continued to associated with the cobble. The presence of cobble resulted in fewer juveniles being captured and a significant increase in the latency until the first juvenile was captured by the predator. Results are discussed with respect to the effects of predation on the distribution and survival of fishes among substrate types.     

Ontogeny of feeding and respiration in larval Atlantic cod Gadus morhua (Teleostei,Gadiformes): I. Morphology

Cranial development in larval Atlantic cod Gadus morhua was studied throughout ontogeny using specimens treated by staining and clearing, scanning electron microscopy and histology. Newly hatched cod larvae have closed mouths, no operculii, five well-developed branchial arches, and transversii ventralis muscles. During the endogenous feeding (yolk-sac) stage, viscerocranial structures remain simple and nonarticulated. Six days after hatching at 5°C, articulation occurs between the quadrate/Meckel's cartilage and the hyomandibula/cranium. Integration of skeletal elements results in a functional jaw that facilitates the transition from endogenous to exogenous feeding. During later ontogenetic stages, the opercular apparatus and levator-operculi coupling develops, facilitating the transition of cutaneous to branchial respiration. Overall, feeding and respiratory needs are met by changes in form (including composition) and function during larval fish growth and are correlated with demands of energy acquisition essential to survival. © 1996 Wiley-Liss, Inc.     

Immunological differences in intestine and rectum of Atlantic cod (Gadus morhua L.)

The defence system of the distal gut (hindgut and rectum) of Atlantic cod, (Gadus morhua L.) was studied using (immuno)histochemical, electron microscopical and real-time quantitative PCR techniques. The uptake and transport of macromolecules in the intestinal epithelium was also investigated.In this study we observed that cod has many and large goblet cells in its intestinal epithelium and that IgM⁺ cells are present in the lamina propria and their number is considerably higher in the rectum than in the intestine. Myeloperoxidase staining revealed low numbers of granulocytes in and under the epithelium of the distal intestine, whereas high numbers were found clustered in the submucosa of the rectum. Electron microscopy not only confirmed these observations, but also revealed the presence of lymphoid cells and macrophages within the intestinal epithelium. Acid phosphatase staining demonstrated more positive macrophage-like cells in the rectum than in the distal intestine. Antigen uptake studies showed a diffused absorption of horse radish peroxidase (HRP) and LTB-GFP, whereas ferritin uptake could not be detected.Basal gene expression of cytokines (IL-1β, IL-8 and IL-10) and immune relevant molecules (hepcidin and BPI/LPB) were compared in both the intestine and rectum and revealed approximately 2–9 times higher expression in the rectum, of which IL-1β expression showed the most prominent difference.The present results clearly indicate that intestinal immunity is very prominent in the rectum of cod.     

Karyotyping and cytogenetic mapping of Atlantic cod (Gadus morhua Linnaeus, 1758)

The Atlantic cod (Gadus morhua) is an important natural resource for northern societies and is now also considered to be a promising candidate for aquaculture. In recent years, much effort has been directed towards the development of genomic tools, and genome initiatives for Atlantic cod have been established. Despite the growing attention devoted to the Atlantic cod genomics, basic aspects of its genome structure and organization remain unknown. Thus, the present work aims to study cytogenetic features of the Atlantic cod as a contribution to the knowledge of this species' genome. The Atlantic cod displays a diploid number of 46 chromosomes, with a karyotypic formula 16 m/sm + 30 st/t. Conventional karyotyping was improved by chromosomal mapping of two classes of repetitive sequences. 18S rDNA clusters were assigned to pairs 2 and 4; small amounts of 18S rDNA clusters were occasionally detected on pair 5. These findings could not be related to the geographical origin of the specimens, but were consistent with the variability of these repeated genes in fish in general. 5S ribosomal gene clusters, apparently corresponding to a single 5S rDNA class, were detected on twelve chromosomes (pairs 11, 12, 14, 17, 20 and 21). The present update of the existing but meagre information on the karyotype of Atlantic cod, plus the first physical mapping of repetitive genes in this species herein, opens the way for an integrated approach that combines genetic and physical mapping with the assembly of the genome of this commercially important species.     

Temperature and sex dependent effects on cardiac mitochondrial metabolism in Atlantic cod (Gadus morhua L.)

To test the hypothesis that impaired mitochondrial respiration limits cardiac performance at warm temperatures, and examine if any effect(s) are sex-related, the consequences of high temperature on cardiac mitochondrial oxidative function were examined in 10 °C acclimated, sexually immature, male and female Atlantic cod. Active (State 3) and uncoupled (States 2 and 4) respiration were measured in isolated ventricular mitochondria at 10, 16, 20, and 24 °C using saturating concentrations of malate and pyruvate, but at a submaximal (physiological) level of ADP (200 µM). In addition, citrate synthase (CS) activity was measured at these temperatures, and mitochondrial respiration and the efficiency of oxidative phosphorylation (P:O ratio) were determined at [ADP] ranging from 25–200 µM at 10 and 20 °C. Cardiac morphometrics and mitochondrial respiration at 10 °C, and the thermal sensitivity of CS activity (Q₁₀=1.51), were all similar between the sexes. State 3 respiration at 200 µM ADP increased gradually in mitochondria from females between 10 and 24 °C (Q₁₀=1.48), but plateaued in males above 16 °C, and this resulted in lower values in males vs. females at 20 and 24 °C. At 10 °C, State 4 was ~10% of State 3 values in both sexes [i.e. a respiratory control ratio (RCR) of ~10] and P:O ratios were approximately 1.5. Between 20 and 24 °C, State 4 increased more than State 3 (by ~70 vs. 14%, respectively), and this decreased RCR to ~7.5. The P:O ratio was not affected by temperature at 200 μM ADP. However, (1) the sensitivity of State 3 respiration to increasing [ADP] (from 25 to 200 μM) was reduced at 20 vs. 10 °C in both sexes (K_m values 105±7 vs. 68±10 μM, respectively); and (2) mitochondria from females had lower P:O values at 25 vs. 100 μM ADP at 20 °C, whereas males showed a similar effect at 10 °C but a much more pronounced effect at 20 °C (P:O 1.05 at 25 μM ADP vs. 1.78 at 100 μM ADP). In summary, our results demonstrate several sex-related differences in ventricular mitochondrial function in Atlantic cod, and suggest that myocardial oxidative function and possibly phosphorylation efficiency may be limited at temperatures of 20 °C or above, particularly in males. These observations could partially explain why cardiac function in Atlantic cod plateaus just below this species׳ critical thermal maximum (~22 °C) and may contribute to yet unidentified sex differences in thermal tolerance and swimming performance.     

Ontogeny of redox regulation in Atlantic cod (Gadus morhua) larvae

The reduction potential of a cell is related to its fate. Proliferating cells are more reduced than those that are differentiating, whereas apoptotic cells are generally the most oxidized. Glutathione is considered the most important cellular redox buffer and the average reduction potential (E_h) of a cell or organism can be calculated from the concentrations of glutathione (GSH) and glutathione disulfide (GSSG). In this study, triplicate groups of cod larvae at various stages of development (3 to 63 days post-hatch; dph) were sampled for analyses of GSSG/2GSH concentrations, together with activities of antioxidant enzymes and expression of genes encoding proteins involved in redox metabolism. The concentration of total GSH (GSH+GSSG) increased from 610±100 to 1260±150 μmol/kg between 7 and 14 dph and was then constant until 49 dph, after which it decreased to 810±100 μmol/kg by 63 dph. The 14- to 49-dph period, when total GSH concentrations were stable, coincides with the proposed period of metamorphosis in cod larvae. The concentration of GSSG comprised approximately 1% of the total GSH concentration and was stable throughout the sampling series. This resulted in a decreasing E_h from −239±1 to −262±7 mV between 7 and 14 dph, after which it remained constant until 63 dph. The changes in GSH and E_h were accompanied by changes in the expression of several genes involved in redox balance and signaling, as well as changes in activities of antioxidant enzymes, with the most dynamic responses occurring in the early phase of cod larval development. It is hypothesized that metamorphosis in cod larvae starts with the onset of mosaic hyperplasia in the skeletal muscle at approximately 20 dph (6.8 mm standard length (SL)) and ends with differentiation of the stomach and disappearance of the larval finfold at 40 to 50 dph (10–15 mm SL). Thus, metamorphosis in cod larvae seems to coincide with high and stable total concentrations of GSH.     

Phenotypic flexibility of digestive system in Atlantic cod (Gadus morhua)

This study examined the restoration of the digestive capacity of Atlantic cod (Gadus morhua Linnaeus) following a long period of food deprivation. Fifty cod (48 cm, 1 kg) were food-deprived for 68 days and then fed in excess with capelin (Mallotus villosus Müller) on alternate days. Ten fish were sampled after 0, 2, 6, 14 and 28 days and the mass of the pyloric caeca, intestine and carcass determined. Two metabolic enzymes (cytochrome c oxidase and citrate synthase) were assayed in white muscle, pyloric caeca and intestine, and trypsin activity was measured in the pyloric caeca. A delay of 14 days was required before body mass started to increase markedly, whereas most of the increase in mass of both the pyloric caeca and intestine relative to fish length occurred earlier in the experiment. By day 14, the activities of trypsin and citrate synthase in the pyloric caeca as well as citrate synthase in the intestine had reached maxima. The growth of the digestive tissues and restoration of their metabolic capacities thus occur early upon refeeding and are likely required for recovery growth to take place. The phenotypic flexibility of the cod digestive system is therefore remarkable: increases in trypsin activity and size of pyloric caeca resulted in a combined 29-fold increase in digestive capacity of the fish during the refeeding period. Our study suggests that Atlantic cod are able to cope with marked fluctuations in food availability in their environment by making a rapid adjustment of their digestive capacity as soon as food availability increases.     

Purification and characterization of pancreatic elastase from Atlantic cod (Gadus morhua)

1. A pancreatic elastase from Atlantic cod (Gadus morhua) has been purified and characterized. 2. The enzyme is a very basic protein with an approximate mol. wt of 28,000. 3. The cod elastase has higher elastin specificity than porcine elastase, and it is inhibited by soybean trypsin inhibitor, which has no effect on porcine elastase. 4. The cod elastase expresses a higher turnover number (kcat) and catalytic efficiency (kcat/Km) than porcine elastase, but it is less thermostable.     

Fertility of gynogenetic Atlantic cod (Gadus morhua L.)

In order to investigate whether meiotic gynogenetic Atlantic cod is fertile and able to produce viable offspring, meiotic gynogenetic females were produced in spring 2010 by activating cod eggs using irradiated sperm. The extrusion of the second polar body was prevented by the application of hydrostatic pressure (56.6 MPa) 36 min after fertilization. In February 2012, their mean round weight was 972 g, and 2580 g in March 2013. In 2012, when the fish were 2 years old, about 52% were mature, 33% were immature, and 13% had undifferentiated gonads. One year later, 77% were mature, 11% were immature, and 11% had undifferentiated gonads. Several of the mature females had malformed gonads, with only one developed ovary lobe or with the two lobes fused. The mean gonadosomic index (GSI) of the 2‐year‐old mature females was 5.2%, with an estimated relative fecundity of 581 000 eggs kg ovary‐free wet weight^?1. Females were stripped for eggs when 2 and 3 years old (2012 and 2013), and fertilized with sperm from normal males. Offspring were obtained from 12 of 17 and 12 of 15 egg batches incubated in 2012 and 2013, respectively, proving that the gynogenetic females are fertile. Furthermore, larvae in all but one of the hatched groups from 2013 had commenced feeding 2 h after being startfed using rotifers 4 days after hatch, indicating viable offspring.     

Allometric growth and condition factor of Atlantic cod (Gadus morhua) fed to satiation: effects of temperature and body weight

The effect of thermal environment on condition factor was examined for six different size-classes of Atlantic cod Gadus morhua fed to satiation. A weight–length relationship for 8 to 1303 g fish reared at 4–20°C indicated an allometric growth ( W  =  aL ^b , a  = 0.0045, b  = 3.257) of cod. Changes in relative condition factor ( K _rel) with temperature were described with a second order polynomial. The most pronounced effect of temperature on body condition was found in the smallest size-classes, but the curves flattened with increased size. Temperature had size-dependent effects on the relative condition factor obtained from an overall weight–length relationship for all fish in the experiment, i.e. K _rel increased with weight at 4°C, but decreased with weight at 16 and 20°C. K _rel remained high for most size-classes at 8 and 12°C. The slopes ( b -values) of the weight–length relationships decreased linearly with temperatures from 4 to 16°C.     

Impacts of regular and random noise on the behaviour,growth and development of larval Atlantic cod (Gadus morhua)

Anthropogenic noise impacts behaviour and physiology in many species, but responses could change with repeat exposures. As repeat exposures can vary in regularity, identifying regimes with less impact is important for regulation. We use a 16-day split-brood experiment to compare effects of regular and random acoustic noise (playbacks of recordings of ships), relative to ambient-noise controls, on behaviour, growth and development of larval Atlantic cod (Gadus morhua). Short-term noise caused startle responses in newly hatched fish, irrespective of rearing noise. Two days of both regular and random noise regimes reduced growth, while regular noise led to faster yolk sac use. After 16 days, growth in all three sound treatments converged, although fish exposed to regular noise had lower body width–length ratios. Larvae with lower body width–length ratios were easier to catch in a predator-avoidance experiment. Our results demonstrate that the timing of acoustic disturbances can impact survival-related measures during development. Much current work focuses on sound levels, but future studies should consider the role of noise regularity and its importance for noise management and mitigation measures.     

Correlations between hemoglobin type and temperature preference of juvenile Atlantic cod Gadus morhua

Atlantic cod (Gadus morhua L.) exhibits polymorphic hemoglobin variants with the HbI locus showing a strong North-South geographic cline in frequency distribution of three main types (1/1, 1/2 and 2/2). This may indicate selective advantages of the different HbI types under various temperature regimes. Despite this only one study has directly examined the temperature preference of the two homozygous types, HbI-1/1 and HbI-2/2, whereas the preference of the heterozygote (HbI-1/2) has never previously been addressed. By exposing fish to a 4–19 °C temperature gradient in an annular preference chamber we recorded the preferred temperature of wild juvenile G. morhua of all three main Hbl types originating from an area where they co-exist. HbI-2/2 G. morhua preferred significantly cooler water (8.9 ± 0.2 °C) compared to the HbI-1/1 group (11 ± 0.6 °C), this difference, however, not being as distinct as previously reported. There was pronounced inter-individual variation in the temperature preference of the HbI-1/2 G. morhua ranging between 6.7 and 13.8 °C, and their overall preference (10.5 ± 0.9 °C) did not differ significantly from either of the homozygous HbI types. Notably, the mean range of utilized temperature (temperature span between 1st and 3rd quartile) was very similar between all 3 Hbl types with 3.2–3.5 °C. Considering the complexity of a trait like temperature preference, there are clearly many other factors besides HbI type that influence the thermal biology of cod, and therefore we also investigated possible associations between genotype and temperature preference for 12 variable candidate gene single nucleotide polymorphisms (SNPs) a priori expected to be related to growth and reproduction. There were, however, no significant correlations between temperature preference and any of the candidate gene SNPs indicating that none of these polymorphisms strongly associates with thermal behavior. Considering however the high-throughput genotyping methods becoming increasingly accessible there is great potential for association studies involving many more genetic markers to identify additional genetic polymorphisms that are important for temperature preference in G. morhua. In conclusion, we support the notion of a ‘warm’ (HbI-2/2) and a ‘cold’ (HbI-1/1) Hb type, although we suggest the difference to be more subtle than previously reported. Furthermore HbI-1/2 G. morhua shows rather inconsistent thermoregulatory behavior. To obtain a more definitive picture of the extent to which thermal niches are realized under natural conditions field observations in areas where the 3 HbI types co-exist should be performed.     

Gut morphology of diploid and triploid Atlantic cod,Gadus morhua

Little is known about the functional consequences of triploidy in Atlantic cod. This study compared the gut morphology of diploid and triploid offspring of wild and selected broodstock. Three‐year‐old triploid offspring of wild cod (mean weight 1695 ± 346 g) had approximately 18% fewer pyloric caeca (125 ± 9 vs 172 ± 14, P < 0.001) and a 23% shorter intestine [Relative Gut Length, (RGL); 1.40 ± 0.17 vs 1.81 ± 0.17, P < 0.05] than their diploid siblings (mean weight: 1820 ± 262 g). Two‐year‐old triploid offspring of selected broodstock (mean weight: 640 ± 64 g) had 20% fewer pyloric caeca (309 ± 17 vs 387 ± 27, P < 0.001) but similar RGL to their diploid siblings (mean weight: 820 ± 69 g). The average number of mucus cells in the columnar epithelium of pyloric caeca was significantly higher in triploid than in diploid cod (54 ± 9 vs 25 ± 5, P < 0.001). There was no correlation between pyloric caeca number and RGL, or between mucus cells and pyloric caeca number, and no significant differences between sexes for any of the measured variables. Overall, the observations highlight some differences in the digestive system of these two ploidy groups that could have an influence on nutrient utilization and performance capacity in triploids compared to diploids.