Study of digestive proteinases and proteinase inhibitors of Daphnia carinata

Quantification of proteases activities and their class structure have been studied in a cladoceran, Daphnia carinata. Protease activity ranged from 0.28 to 0.55 Unit mg⁻¹ protein min⁻¹ with an average value of 0.42±0.06 Unit mg⁻¹ protein min⁻¹. Chymotrypsin activity was more than twofold higher (0.49±0.09 Unit mg⁻¹ protein min⁻¹) than the trypsin activity (0.21±0.02 Unit mg⁻¹ protein min⁻¹). Protease activity and reduction of activity in bands of samples treated with specific inhibitors were documented in photometric assay and substrate SDS–PAGE. Proteinase activity against azocasein was inhibited (91.4±1.5%) with SBTI. PMSF reduced the enzyme activity by 53.1±6.5%, and the azocasein hydrolysis was reduced up to 64.6±3.8% by the specific inhibitor of trypsin, TLCK. In the present investigation, the molecular weight of various activity bands ranged from 16.3 to 51.1 kDa. The molecular weights of several protein bands are similar to protease activity zones. The knowledge of digestive enzyme profiles of fish food organisms generated in the present study may assist in the formulation of age-specific feed.     

Acute effects of benzo[a]pyrene on liver phase I and II enzymes,and DNA damage on sea bream <Emphasis Type="Italic">Sparus aurata</Emphasis>

In the present study biotransformation and detoxification responses to acute exposure to the polycyclic aromatic hydrocarbons benzo[a]pyrene (B[a]P) were investigated in the liver of Sparus aurata (sea bream). Sexually immature gilthead sea bream were treated by intraperitoneal injection of B[a]P (20 mg kg⁻¹) for 6, 12, 24, and 48 h. B[a]P accumulation was quantified in sea bream liver by mean of gas phase chromatography (GPC-MS) after the various exposure periods. The following biological responses were measured: (1) ethoxyresorufin-O-deethylase (EROD) activity, as a phase I biotransformation parameter; (2) liver glutathione S-transferase (GST) activity as a phase II conjugation enzyme. DNA damage was assessed over time using the single-cell gel electrophoresis comet assay. B[a]P bioaccumulation in the liver resulted in a biphasic curve with an increasing uptake up to 5.55 ± 0.67 μg g⁻¹ dry weight after only 6 h exposure and 4.67 ± 0.68 μg g⁻¹ dry weight after 48 h exposure. EROD activity showed a nonsymmetrical bell-shaped kinetic with a maximum at 24 h and lower but significant activities at 12 and 48 h with respect to control animals. Hepatic GST activities were only significant after 48 h exposure. Comet assay showed an increase in liver cells DNA damage with a maximum after 48 h exposure reaching up to 12.17 %DNA in the tail.     

The effects of temperature on the uptake and metabolism of benzo[a]pyrene in isolated gill cells of the gulf toadfish,Opsanus beta

The effects of acclimation temperature and acute temperature change on the uptake and metabolism of the procarcinogen benzo[a]pyrene (BaP) by gill cells of the gulf toadfish, Opsanus beta, were examined. BaP was rapidly accumulated by isolated gill cells and uptake rates were directly proportional to BaP concentration in the medium (1 to 100 μg/ml). Uptake rates were higher in cells isolated from fish acclimated to 18°C when compared to cells from 28°C acclimated fish at all incubation temperatures. When cells were exposed to BaP at the respective acclimation temperatures of the fish, uptake rates were similar (0.14 ± 0.01 at 18°C and 0.12 ± 0.01 μg BaP/s/10 mg cells at 28°C). This finding is discussed in view of results which showed a partial compensation of membrane fluidity in plasma membranes isolated from fish from the two acclimation temperatures. At higher incubation temperatures, cells from fish acclimated to 18°C metabolized BaP at a greater rate than those at 28°C (49.6 ± 1.92 and 43.0 ± 2.24 μg/g/8h, respectively, at 23°C). Low but detectable activities of common biotransformation enzymes (aryl hydrocarbon hydroxylase, glutathione-S-transferase) and cytochrome P-450 content were found, however, no significant differences were evident between cells from fish acclimated to different temperatures. To whom to address correspondence     

Assessment of the Nutritive,Biochemical, Antioxidant and Antibacterial Potential of Eight Tropical Macro algae Along Kachchh Coast,India as Human Food Supplements

Eight tropical marine macro algae were investigated for in-vitro antioxidant, antibacterial, and biochemical properties. The moisture content [% DW (dry weight)] ranged from 7.21 to 14.72%; ash, 24.92 to 47.04%; lipids, 0.73 to 2.67%; protein, 4.56 to 12.59%; and carbohydrate, 30.1 to 48.51%. The % sum of polyunsaturated fatty acids (PUFAs) ranged from 21.71 to 78.22%, while omega-6/omega-3 (ω6/ω3) ratio was from 0 to 2.08, which remained within the prescribed World Health Organization (WHO) standards (<10). The % sum of essential amino acids (EAAs) ranged from 86.18 to 204.66, and Na/K ratio ranged from 0.37 to 2.85. The extracts exhibited significant (P < 0.05) values for total phenol (7.25–16.0 µg PGE mg^–1 DW) and total flavonoid (4.06–15.63 µg QE mg^–1 DW) at 200 µg/ml. Antibacterial activity was tested against the selected food spoilage and pathogenic bacteria. The principal component analysis (PCA) confirmed a positive correlation between total phenolic content, 1, 1-diphenyl-2-picryl-hydrazyl (DPPH) radical-scavenging activities, and ferric reducing antioxidant power (FRAP), which supports the integration of seaweeds as ingredients in functional foods.     

Sperm cryopreservation of silver barb (Barbodes gonionotus): cryoprotectants,cooling rate and storage time on sperm quality

Effectiveness and efficiency of frozen sperm on fertilization and hatching success of eggs from silver barb was examined in relation to cryoprotectants, freezing rate and storage period. Sperm was diluted in calcium‐free Hank's balanced salt solution, equilibrated with dimethylsulphoxide (DMSO), propylene glycol, sucrose or methanol at 5%, 10%, 15% or 20% final concentrations, and frozen in 250‐μL straws using a one‐step freezing procedure (1, 5 and 8°C min^?1 from 25 to ?40°C). Highest post‐thaw sperm motility was found from a treatment using 10% DMSO and 5°C min^?1 (82.2 ± 2.1%), similar to that of 10% DMSO and 8°C min^?1 (87.8 ± 3.2%). Post‐thaw motility of sperm frozen at 5 or 8°C min^?1 was significantly higher than 1°C min^?1. Relative sperm motility declined significantly after 10 months of cryostorage while viability did not change during a 12‐month cryostorage. Average fertilization rates of sperm after 1 and 4 months of storage were 64.5 ± 4.6% and 61.3 ± 3.4%, respectively, similar to those of fresh sperm (69.6–72.3%). Hatching rates of cryopreserved sperm (45.4–51.2%) were similar to those of fresh sperm (51.8–57.8%). This study developed suitable methods for cryopreservation of silver barb sperm that can be used to facilitate hatchery operation.     

Effect of dietary supplementation of periphyton on growth,immune response and metabolic enzyme activities in Penaeus monodon

A 60‐day indoor trial was conducted to study the effect of periphyton supplementation on metabolic and immune responses in tiger shrimp, Penaeus monodon. Periphyton developed over bamboo substrate in outdoor tanks (15 m²) was used as dietary supplement for P. monodon (2.02 ± 0.04 g) reared in 1000 L FRP tanks. Graded levels of periphyton were included in shrimp basal diets: 0% (P0), 3% (P3), 6% (P6), 9% (P9) and P0 diet with natural periphyton (NP) over bamboo substrate. At the end of the trial, P6 and NP showed significantly higher (P < 0.01) growth rate, 23.9% and 20%, respectively, compared with control, P0. Comparatively, lower level of metabolic enzymes, such as lactate dehydrogenase, malate dehydrogenase, aspartate aminotransferase and alanine aminotransferase, was recorded in treatments P3, P6 and NP compared with control, P0. The periphyton‐supplemented group, P3 had significantly higher (P < 0.05) superoxide dismutase (15.83 ± 0.96) and catalase activity (15.73 ± 0.69) compared to 6.88 ± 2.84 and 9.15 ± 0.67 unit mg^?1 protein min^?1_, respectively, in P0. Similarly, higher total haemocyte counts, 32.58 ± 1.30, 28.51 ± 3.12 and 27.26 ± 4.43 × 10⁶ cells mL^?1_, were recorded in P6, NP and P3, respectively, compared to P0, 23.57 ± 1.80 × 10⁶ cells mL^?1. After challenge with Vibrio harveyi, P3 recorded the highest relative percentage survival 67% followed by NP (58%) and P6 (42%) compared with control. However, treatment with highest periphyton inclusion (P9) did not differ significantly with P0 on growth and immunological parameters. This study indicates that periphyton supplementation at 3–6% level improves growth, immune response and metabolic activities in P. monodon.     

Flow field control via aeration adjustment for the enhancement of larval survival of the kelp grouper Epinephelus bruneus (Perciformes:Serranidae)

Flow field control via aeration adjustment for the enhancement of larval survival of the kelp grouper Epinephelus bruneus was examined. Aeration rate of 300 mL min^?1 was introduced during daylight (07:00–19:00 hours) and adjusted to 0, 300 and 900 mL min^?1 at night (19:00–07:00 hours). Larval sinking velocity±SD increased from 0.08 ± 0.05 to 0.26 ± 0.24 cm sec^?1 from 4 to 12 days after hatching (DAH), indicating their susceptibility to sink. Larvae reared in 300 mLmin^?1 attained the highest survival rate at 24.9 ± 3.4%, but remained significantly smaller in growth: 4.54 ± 0.56 mm compared with 4.82 ± 0.53 mm in 900 mL min^?1. The flow field in 300 and 900 mL min^?1 was at 10–20 and 15–25 cm above the bottom of the tank and 8.0 and 1.0 cm beneath the water surface. A favourable rearing condition was observed in 300 mL min^?1 as larvae were away from the bottom and surface areas, thus preventing them from dying due to sinking and surface tension‐related death (STRD). Although sinking death was decreased with an increasing aeration rate, the stronger flow had increased larval susceptibility to STRD. Our findings suggest that aeration at 300 mL min^?1 could enhance larval survival by reducing both sinking death and STRD.     

Leaching of taurine from commercial type aquaculture feeds

Leaching of soluble compounds from pelleted feeds is an issue for the aquaculture industry through increased environmental impact and reduced ingestion essential components. This study was undertaken to examine the leaching rates of taurine, a non‐protein amino acid with critical physiological roles in teleosts. To this end we adapted a new liquid chromatography mass spectrometry (LC‐MS) method for quantifying taurine. Twelve different feeds (4 mm dia.) varying in protein source and taurine levels were examined. Fishmeal content ranged from 0.0% to 45.5% with taurine supplementation ranging from 0.0% to 5.0%. Taurine was extracted and quantified from individual pellets in triplicate at six time points (0, 1, 5, 10, 20 and 40 min). Leaching rates ranged from 0.026 ± 0.005 to 0.826 ± 0.121 mg min^?1 over 40 min at 27°C and were strongly correlated to initial taurine content of the feeds (for distilled water n = 12, P < 0.001, R² = 0.91 for artificial seawater, 25 ppt, n = 4, P = 0.020, R² = 0.96). Loss of taurine from feeds was 59.5 ± 16.5% after 40 min. This study shows that a significant amount of taurine is lost over time from uneaten feed and that taurine supplementation should exceed requirement levels for slow consumers or feed being delivered as multiple additions.     

A comparative study of picomolar affinity 2-[125I]iodomelatonin binding sites in the hearts of three salmonid species

The hearts of three cultured salmonid species, collected at either mid-light or mid-dark were studied for their binding to 2-[¹²⁵I]iodomelatonin, a specific melatonin agonist. The binding was saturable, reversible, and highly specific. The equilibrium dissociation constant (K_d) ranged from 30.1 ± 3.0 pmole 1⁻¹ in Arctic charr (Salvelinus alpinus) to 40.5 ± 2.3 pmole 1⁻¹ in rainbow trout (Oncorhynchus mykiss) indicating a high binding affinity. The maximum density of binding (B_max) was at the low femtomolar level of 0.57 to 0.87 fmole mg⁻¹ protein. Higher B_max appeared to be demonstrated in the mid-light samples when compared to the mid-dark samples but the difference was not significant (p > 0.05). Competition study with various indoles showed the following order of potency: 2-iodomelatonin > melatonin > 6-chloromelatonin ≫ N-acetylserotonin ⋙ serotonin. Guanosine 5′-O-(3-thiotriphosphate) (GTP_γS) strongly inhibited the binding (IC₅₀ = 0.66 μmole 1⁻¹) in the rainbow trout heart, suggesting that these binding sites belong to the superfamily of G-protein linked receptors. Our results suggest the presence of melatonin receptors in the fish heart. In addition, there was no marked intraspecies differences in K_d, B_max and specificity that could be correlated with the phylogeny or life history of the salmonid species.     

Pharmacokinetics and bioavailabilities of 14C-keto-carotenoids, astaxanthin and canthaxanthin, in rainbow trout, Oncorhynchus mykiss

The pharmacokinetics and bioavailabilities of ¹⁴C‐astaxanthin and ¹⁴C‐canthaxanthin were studied in the blood of rainbow trout following intra‐arterial (i.a.) and oral (p.o.) administration. Sixteen months old 1 kg trout were cannulated in the dorsal aorta. [6,7,6′,7′‐¹⁴C]‐keto‐carotenoids were administered i.a. and p.o. at a dose of 573.5 kBq kg^?1 fish body weight for astaxanthin and 836.2 kBq kg^?1 fish body weight for canthaxanthin. After i.a. distribution, total body clearance (Cl_tot) was 17.30±20.29 mL kg^?1 of fish h^?1 for ¹⁴C‐canthaxanthin and 3.30±1.50 mL kg^?1 of fish h^?1 for ¹⁴C‐astaxanthin. The volume of distribution at steady‐state (V_ss) was 208.32±124.79 mL kg^?1 of fish and 71.84±64.15 mL kg^?1 of fish for ¹⁴C‐canthaxanthin and ¹⁴C‐astaxanthin respectively. Less than 0.4% of the administered radioactivity was recovered in urine. Radioactivity (expressed as percent of the dose) excreted in the bile of fish that received ¹⁴C‐canthaxanthin by i.a. route was 20‐fold higher than that observed for fish treated p.o. This ratio was lower for ¹⁴C‐astaxanthin (7.6‐fold). The mean keto‐carotenoid bioavailabilities calculated were 10–15% for both compounds. Findings suggest one daily astaxanthin application is preferable, while 12‐h time intervals between applications are preferable for canthaxanthin.     

The effect of feed supply rate on growth of juvenile perch (Perca fluviatilis)

This study investigates the effect of the feed supply rate within a meal on growth of juvenile perch (Perca fluviatilis). Groups of PIT-tagged, feed-trained juvenile perch (12 g) were held in 100-L tanks at 18 °C, under a 24L:0D photoperiod and fed five meals per day in excess. Feed was applied at rates of 1.2 (Low), 4.2 (Medium) and 14.3 (High) pellets fish⁻¹ min⁻¹ during two periods, each of 50 days. Cumulative meal-time per day ranged from 10 to 225 min. Specific growth rates (SGR) in the two periods were positively correlated, indicating a consistent individual growth performance. There was growth dimorphism between the sexes. Females grew about 20% faster than males, resulting in final mean weights of ≈ 87 and 58 g respectively. Autopsy revealed differences in sex ratios between treatments and the effect of feeding rate was analysed according to sex. There was a tendency for fish fed at the lowest rate to have a higher SGR than fish in other treatments, both for females (L = 1.98 ± 0.0, M = 1.84 ± 0.08, H = 1.88 ± 0.19% bw day⁻¹) and males (L = 1.68 ± 0.01, M = 1.55 ± 0.16, H = 1.57 ± 0.02% bw day⁻¹). Fish fed at the lowest rate also tended to have the lowest feed conversion ratio (L = 0.88 ± 0.05, M = 1.02 ± 0.11, H = 1.04 ± 0.10) but the effects were not significant. These results suggest that juvenile perch can feed efficiently under a range of conditions.     

Aeration rate adjustment at night to prevent sinking syndrome‐related death in the tiger grouper Epinephelus fuscoguttatus (Perciformes:Serranidae) larvae

The effects of different aeration rates at night to prevent sinking syndrome‐related death (SSRD) of the tiger grouper, Epinephelus fuscoguttatus were examined. The aeration rates were fixed at 300 mL min^?1 at daytime (07:00–19:00 hours) and regulated to 0, 300 and 900 mL min^?1 at night (19:00–07:00 hours). Larval survival, growth, feeding intake, sinking velocity, distribution and behaviour, stress level, surface tension‐related death (STRD) and flow velocity distribution were assessed. The occurrence of SSRD in the tiger grouper was observed through the accelerated sinking velocity (Vl) (from 0.15 ± 0.09 cm s^?1 at 4 days AH to 0.41 ± 0.09 cm s^?1 at 12 days AH) coupled with larval passive swimming behaviour at night‐time. On the final day of experiment (15 days AH), larvae reared in 900 mL min^?1 at night had attained significantly higher (P < 0.05) survival (34.4 ± 5.5%), growth (5.8 ± 0.5 mm) and feeding intake (60.46 ± 6.98 ind. larva^?1). A favourable flow field for the tiger grouper was produced in 900 mL min^?1 at night‐time, in which larvae were transported 15–25 cm above the tank bottom and 1.0 cm beneath the water surface. Under these night‐time rearing conditions, larval stress level and number of STRD reared in 900 mL min^?1 compared with those observed in 300 mL min^?1 remained insignificant, indicating that strong turbulence of flow velocity was not detrimental for larvae. Our findings recommend aeration at 900 mL min^?1 at night as this could improve larval survival by reducing SSRD.     

Biochemical and pharmacological characterization of adenosine A1 receptors in eel (Anguilla anguilla) brain

N⁶-cyclohexyl[³H]adenosine ([³H]CHA) was used to label adenosine A1 receptors in membranes prepared from male and female eel whole brain. The A1 receptor agonist [³H]CHA bound saturably, reversibly and with high affinity (K_d = 0.91 ± 0.12 nM; B_max = 120.36 ± 5.2 fmol mg⁻¹ protein). In equilibrium competition experiments, the adenosine agonists and antagonists all displaced [³H]CHA from high-affinity binding sites with the rank order of potency in displacing, characteristics of an A1 adenosine receptor. Mg²⁺ dramatically increased the affinity of [³H]CHA without modifying the maximal binding capacity. The specific binding was inhibited by guanosine 5′-triphosphate (K_i = 2.54 ± 0.98 μM). The [³H]CHA binding sites are ubiquitously distributed with a maximum in cerebellum and a minimum in olfactory bulb. No difference was observed between male and female brain. In eel brain, synaptosomes (P2), stimulation of adenosine 3′,5′-monophosphate (cyclic AMP) accumulation with 10⁻⁵ M forskolin was markedly reduced (45.5%) by treatment with the adenosine A1 receptor agonist CHA (10⁻⁴ M), and the reduction was reversed in presence of the selective A1 receptor antagonist 8-cyclopentyltheophylline (10⁻⁵ M). In superfused eel cerebellar synaptosomes, K⁺ stimulated the release of adenosine in a partially Ca²⁺-dependent manner. The findings, taken together, suggest the hypothesis that adenosine A1 receptors present in eel brain could modulate synaptic transmission, as A1 receptors do in other vertebrates.     

Effect of cryoprotectants, equilibration periods and freezing rates on cryopreservation of spermatozoa of mahseer, Tor khudree (Sykes) and T. putitora (Hamilton)

A study was conducted to standardize a protocol for cryopreservation of spermatozoa of the endangered mahseer, Tor khudree (Sykes) and T. putitora (Hamilton). The suitability of the cryoprotectants, dimethyl sulphoxide (DMSO) and glycerol, and the combination of the two were tested. Four equilibration periods and four freezing rates were also tested for their standardization. A combination of 9% DMSO and 11% glycerol gave significantly higher mean percentage of hatching in both T. khudree (45.59±1.86%) (control 71.08±0.59%) and T. putitora (45.00±1.25%) (control 73.48±1.19%) among the eight different treatments. Among the four different equilibration periods tested, the equilibration period of 30 min^?1 yielded the highest mean hatching percentage in T. khudree (39.46±1.94%) (control 71.70±0.61%) and T. putitora (38.28±1.06%) (control 73.11±0.82%). Freezing straws at a height of 8 cm above LN₂ surface for 10 min^?1 gave higher hatching percentages for both T. khudree (41.75±1.72%) (control 73.99±1.17%) and T. putitora (41.34±2.04%) (control 72.48±1.51%). The study reports the superior performance of the combination of DMSO and glycerol for the first time.     

Cryopreservation of Atlantic cod (Gadus morhua) sperm in large‐volume straws: applications for commercial production and gene banking

In our study, we used a full factorial analysis of variance design to examine the effects of diluent [Mounib's sucrose‐based diluent+hen's egg yolk (EY) and Hanks' balanced salt solution (HBSS)+EY], freezing rate (?2.5, ?5.0 and ?7.5 °C min^?1) and thawing rate (2.5, 5.0 and 7.5 °C min^?1) on motility and velocity of Atlantic cod sperm cryopreserved in 2.5 mL cryogenic straws. We found that post‐thaw sperm performance was strongly influenced by the presence of higher‐order interactions of the factors we tested. For all models broken down by diluent, the 2.5 °C min^?1 thawing rate had the lowest sperm motility recovery index. Mounib's sucrose‐based diluent+EY had the highest motility recovery index at all thawing rates. Mean per cent motility for fresh sperm (87.7±2.9%) was not significantly different than of sperm cryopreserved using Mounib's sucrose‐based diluent+EY, frozen at ?2.5 °C min^?1 and thawed at 5.0 °C min^?1 (77.1±2.9%). For Mounib's sucrose‐based diluent+EY, velocity was significantly higher with sperm thawed at 7.5 °C min^?1, than sperm thawed at 2.5 °C min^?1, while thawing rate had no effect for HBSS+EY. Our findings have implications for cod mariculture and aiding in conservation efforts for a dominant marine fish species.     

Intestinal phytase II: A comparison of activity and in vivo phytate hydrolysis in three teleost species with differing digestive strategies

Phytates are the primary source of phosphorus in animal feeds of plant origin; however, this phytate phosphorus (PP) is only available to growing fish provided it is released from the phytate molecule as inorganic orthophosphate. Two experiments were conducted to compare phytase activity in the intestinal brush border membrane of carnivorous hybrid striped bass Morone chrysops × M. saxatilis, omnivorous tilapia Oreochromis niloticus × O. aureus and omnivorous koi Cyprinus carpio, and to evaluate the capability of these three species for PP digestibility. The results indicate that tilapia exhibit a significantly higher specific intestinal brush border membrane phytase activity of 5 nmol inorganic phosphate released mg⁻¹ protein min⁻¹ than hybrid bass or koi, which do not differ in their lower activities of 3 nmol inorganic phosphate released mg⁻¹ protein min⁻¹. There were no statistical differences among the three species in terms of total intestinal phytase activity, although the level of total intestinal phytase activity exhibited by the two omnivorous species tended to be higher than that of the carnivorous bass. The tilapia were shown to be capable of digesting substantial amounts of dietary PP, with observed apparent PP digestibility values of 50%. The hybrid bass and the koi could digest only 1–2% of the PP present in the diet. It appears that the only species of the three examined for which intestinal phytase activity is of physiologic significance from a practical standpoint of utilizing dietary PP is tilapia. This revised version was published online in July 2006 with corrections to the Cover Date.     

Isolation, partial characterization and localization of integumental peroxidase, a stress-related enzyme in the skin of a teleostean fish (Cyprinus carpio L.)

Biochemical and immunological characteristics of peroxidase activity of the skin epithelium of common carp (Cyprinus carpio) were investigated and compared with peroxidase activity of blood cells. Skin as well as blood-borne peroxidases eluted from the Superdex column as a 135 kDa protein and both probably are tetrameric molecules. Skin peroxidase activity was characterized by a V_max of 51.5 ± 1.3 U mg^-1 min^-1 and a K_M of 1.64 ± 0.18 mM ortho-phenylenediamine (OPD), whereas blood-borne peroxidase was characterized by a 1,000 fold higher specific activity (V_max = 30.5 103 ± 2.5 103 U mg^-1 min^-1) and a higher affinity (K_M = 0.875 ± 0.003 mM OPD). Polyclonal antibodies were raised against concanavalin-A purified skin peroxidase as well as blood-borne peroxidase. Immunocytochemical labelling showed that peroxidase is present in mucous cells and in mucus covering the skin and gill epithelia, as well as in erythrocytes and leucocytes. We conclude that the mucous cells of the skin produce a biochemically distinct peroxidase that is released in the mucus and may contribute to the antimicrobial properties of the mucous layer covering the skin. After exposure of the fish to cadmium the kinetic characteristics of the enzyme activity, as determined in skin homogenates, changed considerably. The V_max increased significantly to 61.9 ± 1.1 U mg^-1 min^-1, and the affinity for OPD increased to the value demonstrated for blood-borne peroxidase (K_M = 0.888 ± 0.045 mM OPD). Increased peroxidase levels after cadmium exposure were also demonstrated immunochemically in a dotblot assay. However, no significant changes were observed when the circulatory system of the fish was perfused prior to sampling, indicating that erythrocytes are a major contributor to the increased peroxidase activity in carp skin during cadmium exposure. This likely reflects the increased vascularization of the connective tissue layer underlying the skin epithelium, which takes place when the fish are exposed to chronic stressors including cadmium. In the cadmium-exposed fish this effect prevented the biochemical detection of stressor-related changes in epithelial peroxidase reported earlier with cytochemical methods.     

Variation in Body Weight and Total Length Among Families of White Bass,Morone chrysops,Fry After Communal Rearing

Variation in body weight and total length among 15 families of Phase I white bass, Morone chrysops, was evaluated in a communal pond. Trait heritabilities (h²) were estimated and family pedigrees were determined a posteriori using microsatellite molecular markers. Fry averaged 36.7 ± 2.6 mm and 0.53 ± 0.10 g across all families after 32 days of communal rearing. The number of offspring identified in our sample per family ranged between three and 28. There were significant differences between families in both body weight and length (P < 0.05). The families clustered into four overlapping groups, but family rankings differed by trait. Both traits showed low heritability (h ² = 0.07, length; h ² = 0.06, weight). Subsequent studies in white bass should include assessment of larger fish to obtain genetic estimates at different stages in the production cycle. This study represents the first effort to assess variation among white bass families for any growth-related trait using a communal rearing approach.     

Mismatch between patterns of circulating and testicular androgens in African catfish, Clarias gariepinus

11-Ketotestosterone (11-KT) is an important plasma androgen in male African catfish. The quantitatively predominating androgen produced by the testis, however, is 11-hydroxyandrostenedione (OHA). Our working hypothesis to explain this mismatch assumed that OHA is converted to 11-KT at extratesticular sites. First, we examined the in vivo metabolism of [³H]-OHA in mature males after sham-operation or removal of either the testes (TC), the seminal vesicles (SVC), or both (TSVC) by analysing the pattern of circulating [³H]-androgens two hours after intravenous injection of [³H]-OHA. [³H]-OHA was converted to [³H]-11-KT to the same extent in all groups, indicating that neither ablation of testes nor of seminal vesicles, or both attenuates this conversion. We then examined the in vitro metabolism of [³H]-OHA by several types of tissues. Liver and seminal vesicle tissue were found to produce significant amounts of [³H]-11-KT. The conversion capacity in vivo was assessed by injecting TSVC-castrated males with increasing doses of radioinert OHA, followed by the quantification of OHA and 11-KT plasma levels. Saturation of the conversion capacity was not reached but the 11-KT production capacity is at least 80 ng per ml of plasma per hour. Moreover, liver fragments were tested for their OHA to 11-KT conversion capacity in vitro using increasing concentrations of radioinert OHA. The 11-KT producing increased with time and OHA concentration. The production rate was 90 pg 11-KT mg^-1 liver h^-1. Considering the results of the surgical experiments and the fact that the total hepatic mass by far exceeds that of the seminal vesicles, we conclude that the hepatic conversion is of primary relevance in vivo.