Swimming performance, oxygen consumption and excess post-exercise oxygen consumption in adult transgenic and ocean-ranched coho salmon

Routine oxygen consumption ( M o ₂) was 35% higher in 1 day starved and 21% higher in 4 day starved adult transgenic coho salmon Oncorhynchus kisutch relative to end of migration ocean-ranched coho salmon. Critical swimming speed ( U _crit) and M o ₂ at U _crit ( M o _2max) were significantly lower in 4 day starved transgenic coho salmon (1·25 BL s⁻¹; 8·79 mg O₂ kg⁻¹ min⁻¹) compared to ocean-ranched coho salmon (1·60 BL s⁻¹; 9·87 mg O₂ kg⁻¹ min⁻¹). Transgenic fish swam energetically less efficiently than ocean-ranched fish, as indicated by a poorer swimming economy at U _crit ( M o _2max     ). Although M o _2max was lower in transgenic coho salmon, the excess post-exercise oxygen consumption (EPOC) measured during the first 20 min of recovery was significantly larger in transgenic coho salmon (44·1 mg O₂ kg⁻¹) compared with ocean-ranched coho salmon (34·2 mg O₂ kg⁻¹), which had a faster rate of recovery.     

Mercury in tigerfish (Hydrocynus vittatus, Castelnau), green happy (Sargochromis codringtonii, Boulenger) and kapenta (Limnothrissa miodon, Boulenger) from Lake Kariba, Zimbabwe

Concentrations of total mercury were determined in Hydrocynus vittatus (Castelnau), Sargochromis codringtonii (Boulenger), and Limnothrissa miodon (Boulenger) from two localities in Lake Kariba, Zimbabwe.
The mean concentrations of total mercury in H. vittatus from Basin 5 and Basin 2 were 0.08 mg kg⁻¹ and 0.094 mg kg⁻¹, respectively. In S. codringtonii, mean concentrations were 0.004 mg kg⁻¹ and 0.026 mg kg⁻¹ for Basins 5 and 2, respectively. No mercury was detected in L. miodon from Basin 2 while samples from Basin 5 had a mean concentration of 0.069 mg kg⁻¹ (wet weight). Total mercury concentrations were also determined on a dry weight basis.
Within each sampling area, total mercury concentrations were significantly different among species ( P  < 0.05). For H. vittatus and S. codringtonii, total mercury concentrations (in the same species) were not significantly different between the two localities ( P  < 0.05).
The factors causing the observed differences in total mercury between similar species from different localities and among different species in the same locality (sampling area) are discussed. From the observed low levels of mercury in all three species, it was concluded that the mercury constituted 'background levels'. These levels are below the maximum concentrations permissible in human fish foods.     

Characterization of the cortisol response following an acute challenge with lipopolysaccharide in yellow perch and the influence of rearing density

Two experiments were performed to characterize the corticosteroid response of yellow perch Perca flavescens following an intraperitoneal injection of lipopolysaccharide (LPS) and determine if sustained differences in rearing density alter this response. In the first experiment, yellow perch were injected with LPS (3 mg kg⁻¹), saline, or handled without receiving any injection. Concentrations of cortisol in plasma were elevated in all groups relative to non‐disturbed fish at 1·5 and 3 h after handling but by 6 h after injection the mean concentration of cortisol in plasma from LPS‐injected yellow perch were three to five times higher than fish before injection and significantly larger than groups of fish not treated with LPS. In the second test, yellow perch were held at different rearing densities (9 v . 18-19 kg m⁻³) for 3, 7 and 14 days before injection with LPS (3 mg kg⁻¹). The cortisol response of yellow perch following LPS injection of fish held for 14 days at high density was significantly lower than that of fish held at the low density for the same duration. Additionally a trend of a decreased cortisol response to LPS injection as duration of holding increased was observed among fish held at high density relative to fish held at low density. These data illustrate that the corticosteroid response of yellow perch following LPS injection is distinct from handling alone and that the magnitude of the response is impacted by rearing density.     

The effect of different dietary iron levels on growth and hepatic iron concentration in juvenile gibel carp (Carassius auratus gibelio)

An 83-day growth trial was conducted using a flow-through system to examine the effects of different dietary iron levels on growth and hepatic iron concentration in juvenile gibel carp ( Carassius auratus gibelio ). Six purified diets supplemented with different levels of iron (0, 10, 30, 60, 100 and 200 mg kg⁻¹) (as ferrous sulfate) were fed to triplicate groups of fish (initial weight 2.12 ± 0.00 g per fish). The results showed that the addition of iron to the basal diet did not significantly affect the specific growth rate (SGR), feed efficiency (FE), survival, red blood cell amount (RBC), hemoglobin content (Hb), mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH) or mean corpuscular hemoglobin concentration (MCHC). Hepatic iron concentration and hematocrit (Hct) were significantly influenced by dietary iron level (P < 0.05). On the basis of the iron concentration for the maintenance of optimum hepatic iron concentration and Hct, it was concluded that the dietary iron concentration of juvenile gibel carp should be not less than 202 mg Fe kg⁻¹ diet.     

The compatibility of the fungicide azoxystrobin with Pochonia chlamydosporia, a biological control agent for potato cyst nematodes (Globodera spp.)

The biological control potential of Pochonia chlamydosporia against root-knot ( Meloidogyne spp.) and cyst-forming ( Heterodera and Globodera spp.) nematodes is widely appreciated. In spite of this, little has been undertaken to determine the compatibility of this fungus with modern fungicides. A series of experiments were undertaken to investigate the effect of azoxystrobin on P. chlamydosporia . Initial Petri dish experiments found a significant reduction in the growth of P. chlamydosporia . EC₅₀ values were calculated at 7 and 14 mg L⁻¹, respectively, for hyphal growth and chlamydospore germination. A microcosm experiment based on counts of the colony-forming units of the re-isolated fungus showed a reduction in EC₅₀ values for azoxystrobin in soil after 12, 20 and 40 days incubation (1.25, 1.00 and 3.00 mg kg⁻¹ soil, respectively). There was evidence the fungus recovered over time in response to azoxystrobin application. This was also demonstrated in a glasshouse experiment where EC₅₀ values of 1.32 and 4.4 mg kg⁻¹ soil were obtained for 35 and 49 days after planting (DAP), respectively.     

Effect of Lactobacillus gasseri BNR17 on blood glucose levels and body weight in a mouse model of type 2 diabetes

Aims:  To investigate the effect of Lactobacillus gasseri BNR17 isolated from human breast milk on blood glucose and body weight in type 2 diabetic animals.
Methods and results:  db/db mice were divided into one control group and five sample groups; the sample groups received BNR17 (10⁷, 10⁸, 10⁹ and 10¹⁰ CFU) or rosiglitazone (8 mg kg⁻¹) orally twice a day for 12 weeks. BNR17 groups had a dose-dependent reduction in food, water intake and amount of excrement. Body weight loss was not seen in the BNR17 groups. Fasting and postprandial 2 h blood glucose levels were significantly lower in the BNR17 (10¹⁰ CFU) group compared with the control group. HbA1c decreased in the BNR17 group, although it was not statistically significant. During the oral glucose tolerance test, the BNR17 groups exhibited dose-dependent improvement in glucose sensitivity.
Conclusions:  Lactobacillus gasseri BNR17 has a suppressing effect on blood glucose levels and improved diabetic symptoms in db/db mice.
Significance and Impact of the Study:  Blood glucose-lowering lactic acid bacteria are expected to be useful as a therapeutic for treating type 2 diabetes in humans.     

Stimulation of biomethanation by Clostridium sp. PXYL1 in coculture with a Methanosarcina strain PMET1 at psychrophilic temperatures

Aim:  Bioaugumentation of low temperature biogas production was attempted by addition of cold-adapted Clostridium and a methanogen.
Methods and Results:  A psychrotrophic xylanolytic acetogenic strain Clostridium sp. PXYL1 growing optimally at 20°C and pH 5·3 and a Methanosarcina strain, PMET1, growing optimally on acetate and producing methane at 15°C were isolated from a cattle manure digester. Anaerobic conversion of xylose at 15°C with the coculture of the two strains was performed, and batch culture methane production characteristics indicated that methanogenesis occurred via acetate through 'acetoclastic' pathway. Stimulation studies were also undertaken to evaluate the effect of exogenous addition of the coculture on biogas yields at 15°C. Addition of 3 ml of PXYL1 at the rate of 12 × 10² CFU ml⁻¹ increased the biogas 1·7-fold (33 l per kg cowdung) when compared to control (19·3 l per kg cowdung) as well as increased the volatile fatty acid (VFA) levels to 3210 mg l⁻¹ when compared to 1140 mg l⁻¹ in controls. Exogenous of addition of 10 ml PMET1 inoculum at the rate of 6·8 ± 10² CFU ml⁻¹ in addition to PXYL1 served to further improve the biogas yields to 46 l kg⁻¹ as well as significantly brought down the VFA levels to 1350 mg l⁻¹.
Conclusions:  Our results suggest that the rate-limiting methanogenic step at low temperatures could be overcome and that biogas yields improved by manipulating the population of the acetoclastic methanogens.
Significance and Impact of the Study:  Stimulation of biomethanation at low temperature by coculture.     

Endurance swimming of European eel

A long‐term swim trial was performed with five female silver eels Anguilla anguilla of 0·8–1·0 kg ( c . 80 cm total length, L _T) swimming at 0·5 body lengths (BL) s⁻¹, corresponding to the mean swimming speed during spawning migration. The design of the Blazka‐type swim tunnel was significantly improved, and for the first time the flow pattern of a swim tunnel for fish was evaluated with the Laser‐Doppler method. The velocity profile over three different cross‐sections was determined. It was observed that 80% of the water velocity drop‐off occurred over a boundary layer of 20 mm. Therefore, swim velocity errors were negligible as the eels always swam outside this layer. The fish were able to swim continuously day and night during a period of 3 months in the swim tunnel through which fresh water at 19° C was passed. The oxygen consumption rates remained stable at 36·9 ± 2·9 mg O₂ kg⁻¹ h⁻¹ over the 3 months swimming period for all tested eels. The mean cost of transportation was 28·2 mg O₂ kg⁻¹ km⁻¹. From the total energy consumption the calculated decline in fat content was 30%. When extrapolating to 6000 km this would have been 60%, leaving only 40% of the total energy reserves for reproduction after arriving at the spawning site. Therefore low cost of transport combined with high fat content are crucial for the capacity of the eel to cross the Atlantic Ocean and reproduce.     

Farm-scale trials to compare the entomopathogenic fungus Beauveria bassiana with pirimiphos methyl + deltamethrin and essential oil of lemon grass for protection of stored cowpea against Callosobruchus maculatus (Coleoptera: Bruchidae)

In trials conducted in Benin, conidia of Beauveria bassiana were evaluated as a control method against the cowpea weevil, Callosobruchus maculatus , in stored cowpea. In the first trial using a high artificial infestation of C.   maculatus in 8-kg batches of cowpea in jerry cans under typical conditions, concentrations of 1 × 10⁹ and 1 × 10¹¹ conidia kg⁻¹ were compared with lemon grass oil at 2 mL kg⁻¹ and the synthetic pesticide mixture of 1.5% pirimiphos methyl + 0.05% deltamethrin at 0.5 g kg⁻¹. After 2 months of storage, seed losses (SD) were 20.63 (5.3)% in the untreated control, 8.04 (3.2)% in the low-dose B.   bassiana group, 3.12 (1.3)% in the high-dose B.   bassiana group, 2.52 (0.4)% in the lemon grass oil group and 0% in the pirimiphos methyl + deltamethrin group. In a second trial with natural infestations in 50-kg batches of cowpea in 200-L drums, treatment with B.   bassiana 1 × 10¹¹ conidia kg⁻¹ was compared with pirimiphos methyl + deltamethrin at 0.5 g kg⁻¹. After 6 months of storage on six farms, losses reached 30.76 (1.5)% in the control, 1.28 (0.2)% in the pirimiphos methyl + deltamethrin group and 3.69 (0.6)% in the B.   bassiana group.     

Automated sperm morphology analysis in fishes: the effect of mercury on goldfish sperm

This study is the first to examine the morphology of fish sperm using automated sperm morphology analysis (ASMA). The technique was applied to investigate the effect of an environmental pollutant, mercury, on the sperm morphology of goldfish Carassius auratus , and the effects on sperm morphology were compared with those on sperm motility. Goldfish sperm flagellar length was significantly shortened after instant exposure to 100 mg l⁻¹ (368 µM) mercuric chloride, while curvilinear velocity (VCL) and the percentage of motile sperm were significantly decreased at mercuric chloride concentrations of 1 and 10 mg l⁻¹ (3·68 and 36·8 µM), respectively. After 24 h exposure to 0·001 mg l⁻¹ (0·0037 µM) mercuric chloride, flagellar length was significantly reduced in 38% of the spermatozoa. Following exposure to 0·1 mg l⁻¹ (0·37 µM) mercuric chloride for 24 h, however, the majority of spermatozoa (98%), had significantly shortened flagella and increased sperm head length, width and area. Sperm motility was also significantly decreased at 0·1 mg l⁻¹ (0·37 µM) mercuric chloride, probably due to the significantly reduced flagellar length at this concentration. This study shows that the morphological examination of fish sperm by ASMA provides, not only, an excellent tool for monitoring reproductive disruption caused by environmental pollution, but also has applications to other areas of fish reproductive biology, such as cryopreservation and aquaculture.     

Cellular and biochemical response to Cr(VI) in Stenotrophomonas sp.

Chromium (Cr)-resistant bacteria isolated from a soil with 6 g kg⁻¹ of Cr were identified based on 16S rRNA gene sequence analysis as a Stenotrophomonas , and designated as JD1. Growth of JD1 was accompanied by transformation of Cr(VI) to Cr(III) in liquid medium initially containing 300 mg L⁻¹ Cr(VI), the maximum concentration allowing growth. JD1 produced the highest levels of a Cr(VI)-binding exopolysaccharide when grown in medium with 100 mg L⁻¹ Cr(VI). The relative exopolysaccharide monosaccharide composition was analysed by HPLC, which showed that rhamnose+galactose was the major component, and that its relative level increased when cells were grown with Cr(VI). JD1 grew as a biofilm on various inert surfaces. Biofilm macromolecular composition analysis indicated that the relative levels of exopolysaccharide and protein were more abundant in biofilms grown in 100 mg L⁻¹ Cr(VI), whereas relative uronic acid levels remained constant. Biofilm cells exposed to Cr(VI) were elongated, grouped in clusters and exopolysaccharide obtained from the biofilm extracellular matrix had an enhanced capacity to bind Cr(VI). Exopolysaccharide production and composition, and biofilm growth are discussed as a mechanism of protection that allows survival during Cr(VI) stress.     

Growth performance, mortality and carotenoid pigmentation of fry offspring as affected by dietary supplementation of astaxanthin to female rainbow trout (Oncorhynchus mykiss) broodstock

Growth performance, mortality and carotenoid pigmentation were studied in triplicate groups each with 1000 swim-up larvae of rainbow trout ( Oncorhynchus mykiss ), derived from five groups of female broodstock fed diets with 0.07, 12.5, 33.3, 65.1 or 92.9 mg astaxanthin kg⁻¹, respectively. The first feeding fry (initial weight range from 113 to 148 mg) were fed a diet not supplemented with carotenoids in an experiment lasting 45 days. Fry were initially sampled for astaxanthin content and initial weight, and in subsequent 15-day intervals to determine weights, condition factors (CF), specific growth rates (SGR) and thermal growth coefficients (TGC). Total carotenoid concentration of the larvae was highly linearly correlated to that of the eggs ( r ² = 0.97, P = 0.002). About 59–67% of fry carotenoids consisted of esterified astaxanthin, and on average 39.7% of the egg carotenoids were recovered in the fry. Overall (0–45 days) SGRs and TGCs were significantly higher (P < 0.05) in the offspring of the four groups of females fed supplemented diets (12.5–92.9 mg astaxanthin kg⁻¹) than in offspring of females fed the non-supplemented diet. TGCs (0–45 days) within groups derived from broodstock supplemented with astaxanthin were similar (P > 0.05), but higher than in the group derived from females fed the diet not supplemented with astaxanthin (P < 0.05). Mortality (average 0.76%) was not significantly affected by treatment. The study indicates that dietary supplement of astaxanthin (>12.5 mg kg⁻¹) to maternal broodstock diets improves offspring SGR and TGC with up to 33 and 38%, respectively.     

Geophagy and nutrient supplementation in the Ngorongoro Conservation Area, Tanzania, with particular reference to selenium, cobalt and molybdenum

Wildlife and livestock in the Ngorongoro Conservation Area (NCA), Tanzania, commonly practise geophagy. We investigated the nutrient content of nine earth licks and adjacent topsoils in the NCA, and for comparative purposes, two salt pans in north-eastern Tanzania, licks in central Tanzania and a lick near Puerto Maldonado, Peru. The licks had no consistent pattern of nutrient enrichment relative to unutilized topsoils, although the Na content of licks was greater than that of the adjacent topsoils at all but one site. The three largest licks in the NCA (Gibbs Farm, Seneto and Ascent Road) were enriched relative to topsoils and global averages in Se (maximum of 4.7 mg kg⁻¹), Co (maximum of 107 mg kg⁻¹) and/or Mo (maximum of 7.4 mg kg⁻¹). We suggest that licks do provide supplemental Na, but that Se, Co and/or Mo at the largest licks provide easily overlooked nutritional benefits and are perhaps the primary target for geophagy at these sites. Subsoil zones of clay deposition are likely to be enriched in various elements through illuviation or mineral precipitation from solution. Animals may use the taste of NaCl as a clue for locating such zones where they are likely to find a greater quantity of micronutrients relative to other soils. These findings have consequences for conservation and pastoralism in that these large licks may be key resources, providing micronutrients that are essential for maintaining the health and fecundity of animal as well as human populations in the region.     

Effects of osmolality and ions on the motility of stripped and testicular sperm of freshwater-and seawater-acclimated tilapia, Oreochromis mossambicus

A significantly higher concentration of testicular spermatozoa was obtained from freshwater Oreochromis mossambicus (9·9×10⁹ spermatozoa ml⁻¹) than seawater O. mossambicus (4·6×10⁹ spermatozoa ml⁻¹). The mean osmolality of the urine of freshwater fish (78·5 mOsmol kg⁻¹) was significantly different from that of seawater fish (304·8 mOsmol kg⁻¹). The mean length of the mid-piece of the spermatozoa together with the tail was more variable in freshwater O. mossambicus (8·80±0·23μm) than in seawater specimens (8·27±0·18 μm). Stripped sperm of freshwater O. mossambicus was highly contaminated by urine which was a good activator of sperm motility in O. mossambicus held in both fresh and sea water. The osmolality for initiation of motility in freshwater O. mossambicus spermatozoa was from 0 to 333 mOsmol kg⁻¹ while for seawater O. mossambicus spermatozoa it was from 0 to 1022 mOsmol kg⁻¹. The optimum osmolality for motility was from 70 to 333 mOsmol kg⁻¹ for freshwater O. mossambicus spermatozoa and from 333 to 645 mOsmol kg⁻¹ for seawater fish. In freshwater O. mossambicus spermatozoa, the presence of 20 mM CaCl₂ increased the permissive osmolality of NaCl from 184 to 645 mOsmol kg⁻¹. For seawater O. mossambicus spermatozoa, solutions of NaCl devoid of CaCl₂ were unable initiate motility, but the addition of 1·5 to 30 mM CaCl₂ to the NaCl solution (0–934 mOsmol kg₁) had a full motility initiating effect.     

Constitutive mineralization of low concentrations of the herbicide linuron by a Variovorax sp. strain

The mineralization of the herbicide linuron at concentrations of μg and mg L⁻¹ was studied in liquid batch experiments with Variovorax sp. strain SRS16. The strain was highly efficient at mineralizing a range of linuron concentrations (0.002–10 mg L⁻¹) with 20–60% of the added ¹⁴C-ring-labeled linuron metabolized to ¹⁴CO₂ within hours to days depending on the initial linuron concentration and incubation period. At mg L⁻¹ linuron concentrations the mineralization activity by SRS16 was inducible and a shift to constitutive mineralization activity was apparent with a reduction in the linuron concentration to μg L⁻¹ levels. This study revealed that strain SRS16 is a promising candidate for bioaugmentation of water or soil resources contaminated with low linuron concentrations.     

Long-term ammonia exposure of turbot: effects on plasma parameters

Turbot juveniles were exposed to four ammonia concentrations [0·17 (L), 0·34 (M), 0·73 (MH) and 0·88 (H) mg l⁻¹ NH₃-N] for different exposure durations (28 days minimum to 84 days). Their physiological status and growth performances were compared to a control group [0·004 (C) mg l⁻¹ NH₃-N]. No growth was observed in the H group, and by day 57, mass increase in the MH group was only 15% of that in group C. During the first month growth in the L group was similar to that in control group while it was lower (33%) in the M group; afterwards the L and M groups had a similar growth (half that of controls). Accumulation of total ammonia nitrogen (TA-N) in plasma was dependent on ambient ammonia concentrations. Plasma urea levels in ammonia-exposed fish were lower, similar or greater than in controls (depending on ammonia concentration or exposure duration). Osmolarity, Cl^– and Na⁺ plasma concentrations were stable in the L and M groups. The increases in Na⁺, Cl^–, K⁺ and total Ca concentrations observed by the end of the experiment in the H and MH groups suggest that fish failed to adapt. There was an initial rise in plasma cortisol in all ammonia-exposed groups followed by a return to basal level (1·7–4 ng ml⁻¹) in the L and M groups. In group MH, plasma cortisol peaked at 42 ng ml⁻¹ by day 14, and after a decline at c . 1 month (14 ng ml⁻¹), it rose again.     

Potential of Bacillus sp. to produce polyhydroxybutyrate from biowaste

Aim:  To test the Bacillus strains for their abilities to produce polyhydroxybutyrate (PHB) from different sugars and biowaste (Pea-shells).
Methods and Results:  Six Bacillus strains were checked for their ability to produce PHB from GM2 medium supplemented with different sugars at the rate of 1% (w/v) and from biowaste and GM2 (BW : M) combinations (3 : 7, 1 : 1, 7 : 3). Glucose supplemented GM2 medium resulted in maximum PHB production of 435 mg l⁻¹ constituting 31–62% w/w of the total cell dry mass. Substituting GM2 medium to the extent of 50% with biowaste (pea-shell slurry) resulted in 945–1205 mg l⁻¹ PHB (55–65% w/w). Optimization for additional nitrogen supplementation, inoculum size resulted in a final PHB production of 3010–3370 mg l⁻¹ equivalent to 300 g kg⁻¹ biowaste (dry wt).
Conclusion:  The Bacillus strains were able to produce PHB from biowaste (Pea-shells) as cheap source of substrate.
Significance and Impact of the Study:  This is the first report on usage of pea-shells as feed for PHB production, opening new possibilities for its use for production of PHB and Bacillus as potential candidate for the purpose.     

Fungal endophytes from Acer ginnala Maxim: isolation, identification and their yield of gallic acid

Aims:  The aim of the study was to isolate the endophytic fungi from Acer ginnala and screen isolates rich in gallic acid.
Methods and Results:  After epiphytic sterilization, 145 fungal endophytes were isolated from the stem, annual twig and seed of Acer ginnala . The endophytes were grouped into ten different taxa, Phomopsis sp., Neurospora sp., Phoma sp., Epicoccum sp., Penicillium sp., Alternaria sp., Fusarium sp., Trichoderma sp., Cladosporium sp. and a species of Pleosporales Incertae Sedis , by their morphological traits and ITS-rDNA sequence analysis. The content and yield of gallic acid of 141 isolates were determined by HPLC. On average, the species of Pleosporales Incertae Sedis had the highest content and yield of gallic acid (13·28 mg g⁻¹ DW; 119·62 mg l⁻¹), while Alternaria sp. had the lowest.
Conclusions:  Of 141 fungal endophytes from A. ginnala , Phomopsis sp. isolate SX10 showed both the highest content and the highest yield of gallic acid (29·25 mg g⁻¹ DW; 200·47 mg l⁻¹).
Significance and Impact of the Study:  Endophytic fungi isolated from A. ginnala may be used as potential producers of gallic acid and other compounds with biological activities, or functioned as elicitors to produce natural compounds.     

Sulphur amino acid requirement of juvenile Asian sea bass Lates calcarifer

The dietary requirement of juvenile Asian sea bass Lates calcarifer Bloch for total sulphur amino acids was studied. Fish (average initial weight of 2.59 ± 0.08 g) were reared in twelve 500 L fibreglass tanks provided with flow-through seawater at 26°C and salinity of 31 p.p.t. for 12 weeks. They were fed semi-purified test diets containing 6.2, 7.2, 8.1, 9.0, 10.8, or 12.6 g methionine kg⁻¹ dry diet and a basal level of 3.1 g cystine kg⁻¹ dry diet. The mean crude protein of the diets (containing defatted Peruvian fishmeal, squid meal, soybean meal, and free amino acid mixture to simulate the pattern of hydrolysed sea bass protein) was 46.02%. The crude fat content of the diets was 10.51% from a 1 : 1 mixture of cod liver oil and soybean oil. Survival was 100% in all treatments. On the basis of the growth response, the total sulphur amino acid requirement of juvenile Asian sea bass was estimated to be 13.4 g kg⁻¹ dry diet (2.9% of protein). Fish fed low levels of l -methionine had significantly lower weight gains and feed efficiency ratios as well as slightly higher hepatosomatic indices. No nutritional deficiency signs were observed other than growth depression in fish fed on diets that were low in methionine. This information is valuable in further refinement of formulations of practical diets for the Asian sea bass.     

Batch marking of otoliths and fin spines to assess the stock enhancement of Argyrosomus japonicus

Juvenile mulloway Argyrosomus japonicus (54·6 ± 4·6 mm total length, mean ±  s . e .) were immersed in a range of oxytetracycline (OTC) solutions ranging between 0–600 mg l⁻¹ in salinities of 5 (diluted sea water) and 35 (undiluted sea water), and alizarin complexone (ALC) solutions ranging between 0–60 mg l⁻¹ in undiluted sea water, for 6, 12 and 24 h. Optimal marking conditions were 600 mg l⁻¹ OTC for 24 h in a salinity of 5, and 30 mg l⁻¹ ALC for 12 h respectively. Mark quality (MQ) was assessed using a score of 0–3 in both otoliths and anal fin spines, with a score >2 found to be acceptable for adequate mark identification. Acceptable marks were not produced using OTC in undiluted sea water. Immersion in OTC or ALC, or reduced salinity had no effect on survival relative to controls. Transverse sections of vertebrae from the ALC and OTC treatments with the highest otolith mark quality showed no discrete marks. Optimal marking techniques were used to produce double marks with a 3 day interval between marking, and marking techniques were applied to 130 000 juvenile mulloway in batch mode with minimal mortality. A numerical model of the chemical behaviour of OTC in sea water describes the decline of available OTC in increasing salinity, so that a species' salinity tolerance and successful marking can be optimized.