Occurrence of Listonella anguillarum in seed production environments of Japanese flounder Paralichthys olivaceus (Temminck et Schlegel)

The present study was undertaken to investigate the distribution of Listonella anguillarum in the rearing water, fish and diets (rotifers) of Japanese flounder (Paralichthys olivaceus). A total of 793 isolates were obtained from the seed production environment of Japanese flounder and 175 out of them were identified as L. anguillarum by biochemical characterization, polymerase chain reaction (PCR) detection for VAH1 haemolysin gene and phylogenetic analysis of 16S ribosomal deoxyribonucleic acids (rDNA) sequences. These results strongly suggested that L. anguillarum is rapidly and accurately identified by the combination of incubation on thiosulphate–citrate–bile salt–sucrose agar at 35°C overnight and PCR detection for the VAH1 haemolysin gene. All flounder specimens and all rotifer samples harboured L. anguillarum at high densities of 6.9 × 10³–6.3 × 10⁵ colony forming units (CFU) g^?1 and 1.5 × 10⁴–2.3 × 10⁶ CFU g^?1, respectively, while as low as 5.0 × 10⁰–2.0 × 10¹ CFU mL^?1 of L. aguillarum were detected in only two of 11 seawater samples, even though no vibriosis occurred in larval and juvenile flounder of tanks. This fact strongly suggests that L. anguillarum is an inhabitant in the seed production environments of Japanese flounder.     

Bacterial flora associated with the giant freshwater prawn Macrobrachium rosenbergii, in the hatchery system

The objective of this study was to understand the microbial flora associated with the hatchery system of giant fresh water prawn, Macrobrachium rosenbergii during an entire rearing cycle. Bacteriological and physico-chemical analysis was done for different samples of water, larvae, and Artemia. The total bacterial load in well water, seawater and inlet water varied from 10¹ to 10⁵ cfu ml^− 1 with higher counts seen in larval rearing tank (LRT) water. The Vibrio count ranged between 10¹ to 10³ cfu ml^− 1. Larval samples harboured a bacterial load of 10⁶ to 10⁷ cfu/10 larvae. The bacterial load in Artemia hatching water ranged from 4.90 × 10⁴ to 5.63 × 10⁶ cfu ml^− 1 while Artemia had a load ranging from 1.08 × 10⁷ to 2.09 × 10⁹ cfu g^− 1. Vibrio count in the LRT water ranged from 10¹-10³ cfu ml^− 1 while the count in larvae ranged from 10² to 10⁴ cfu/10 larvae. The bacterial genera were predominantly Gram-negative and comprised of Aeromonas spp., Pseudomonas spp., Vibrio spp. and Bacillus spp. and non-spore formers (NSF) were the dominant Gram-positive bacteria. This study documents the bacterial flora associated with Macrobrachium hatchery system during a regular normal run. Knowledge of the qualitative and quantitative aspects of bacterial flora in the hatchery would help to understand disturbances, if any, brought about during disease outbreaks.     

Growth, survival and biochemical composition of spider crab Maja brachydactyla (Balss, 1922) (Decapoda: Majidae) larvae reared under different stocking densities, prey:larva ratios and diets

The spider crab Maja brachydactyla is overexploited on the NW coast of Spain. Aquaculture of this species can be the solution to the problem, and consequently, several attempts of intensive larval rearing have been conducted. However, most of the studies already published do not provide enough zoo technical data, especially in terms of larval and prey densities or the nutritional quality of diets used for rearing.Three experiments were carried out to evaluate the conditions for intensive larval rearing of M. brachydactyla. Larval stocking density (10, 50 and 100 larvae L^− 1), prey:larva ratio (15, 30 and 60 enriched Artemia larva^− 1) and diet (enriched Artemia, non-enriched Artemia and polychaete supplement) effects on growth and survival of this species were studied. For larval culture nine, 35 L, 150 μm mesh-bottomed PVC cylinders (triplicates for each treatment and larval stage) connected to a recirculation unit, were used. Temperature and salinity were kept constant at 18 °C and 36‰ respectively. A 12 to 18 day trial was conducted for each experiment and samples of larvae were collected at each larval stage (zoea I, zoea II, megalopa) in the inter-molt phase and at first juvenile. Survival, carapace length and width, dry weight (DW), and proximate biochemical content (protein, carbohydrates and total lipid) as well as lipid class composition were determined.Stocking densities of 100 larvae L^− 1 resulted in higher growth in DW and higher content in lipids and protein for zoea I (ZI) and zoea II (ZII) than 10 larvae L^− 1. However, survival decreased with increasing stocking density.The use of 60 preys larva^− 1 produced larvae with significantly higher DW and protein content, especially at ZII stage, than lower prey densities. Survival rate obtained feeding 60 preys larva^− 1 up to the megalopa stage was almost two-fold (42.2%) the rate obtained using 15 preys larva^− 1 (24.8%).Larvae fed on enriched Artemia (EA) showed an increase in weight up to megalopa (518.9 ± 26.5 μg) in contrast to larvae fed on non-enriched prey (A) (467.9 ± 6.9 μg). Variation in DW correlated with the total lipid content (L) of the larvae (L_EA = 70.1 ± 37.5 μg ind^− 1; L_A = 28.9 ± 3.2 μg ind^− 1) especially in terms of neutral lipids. The use of an initial density of at least 50 larvae L^− 1 and 60 enriched Artemia larva^− 1 can be considered the most adequate rearing parameters in order to obtain good results in growth and survival of M. brachydactyla.     

The tetrasporophyte of Asparagopsis armata as a novel seaweed biofilter

The red seaweed Asparagopsis armata (Harvey; Rhodophytae, Bonnemaisoniaceae) produces biologically active secondary metabolites that are valuable natural ingredients for cosmetics and medicine and its cultivation may therefore be a profitable venture. The tetrasporophyte of this species (“Falkenbergia rufolanosa”) was successfully tank-cultivated as a continuous biofilter for the effluent of a commercial fish farm in southern Portugal. Optimal stocking density for highest biomass yield and a low level of other algal species in winter and late spring was 5×g centrifuged fresh weight l^− 1. The effect of total ammonia nitrogen supply (TAN flux) on biofiltration and biomass yield was investigated in winter and spring. Results revealed that A. armata is currently the seaweed-biofilter with the highest TAN removal of up to 90 μmol l^− 1 h^− 1 at a TAN flux of about 500 μmol l^− 1 h^− 1. In the tanks used, this is equivalent to a removal of up to 14.5 g TAN m^− 2 day^− 1. At a lower TAN flux of about 40 μmol l^− 1 h^− 1, TAN removal by A. armata is more than double to what is reported at this flux for another successful seaweed biofilter, the genus Ulva. Monthly variation of A. armata biomass yield peaked in May and was lowest in January. At TAN fluxes between 300 and 400 μmol l^− 1 h^− 1, an average water temperature of 21.7 °C and a total daily photon flux density of 47 Mol m^− 2, seaweed yield was over 100 g DW m^− 2 day^− 1 with a recorded maximum of 119 g. During spring, autumn and early summer, the biomass of A. armata within the experimental tanks doubled every week. A model for the up scaling of this finfish integrated aquaculture of A. armata varies the investment in biofilter surface area and estimates the return in biofiltration and biomass yield. Highest TAN removal efficiencies will only be possible at low TAN fluxes and a very large biofilter area, resulting in a low production of biomass per unit area. To remove 50% of TAN from the effluent (1 mt Sparus aurata; 21 °C), 28 m² of biofilter, designed to support a water turnover rate of 0.8 Vol h^− 1 would be necessary. This system produces 6.1 kg FW (1.5 kg DW) of A. armata per day and has the potential to turn biofiltration into an economically sustained, beneficial side effect.     

In situ studies on the bioaccumulation of microcystins in the phytoplanktivorous silver carp (Hypophthalmichthys molitrix) stocked in Lake Taihu with dense toxic Microcystis blooms

The phytoplanktivorous silver carp is an important biomanipulation fish to control cyanobacterial blooms and is also a food fish with the greatest production in China. The accumulation of the hepatotoxic microcystins (MCs) determined by LC-MS in various organs of silver carp was studied monthly in Lake Taihu dominated by toxic Microcystis aeruginosa. Average recoveries of spiked fish samples were 78% for MC-RR and 81% for MC-LR. The highest content of MCs was found in the intestine (97.48 μg g^− 1 DW), followed by liver (6.84 μg g^− 1 DW), kidney (4.88 μg g^− 1 DW) and blood (1.54 μg g^− 1 DW), and the annual mean MC content was in the order of intestine > liver > kidney > blood > muscle > spleen > gallbladder > gill. Silver carp could effectively ingest toxic Microcystis cells (up to 84.4% of total phytoplankton in gut contents), but showed fast growth (from 141 g to 1759 g in 1 year in mean weight). Silver carp accumulated less microcystins in liver than other animals in the same site or other fish from different water bodies at similar level of toxin ingestion. There was possible inhibition of the transportation of the most toxic MC-LR across the gutwall. Muscle of silver carp in Lake Taihu should not be consumed during period of dense Microcystis blooms while viscera were risky for consumption in more months.     

The influence of stocking density, light and temperature on the growth, production and nutrient removal capacity of Porphyra dioica (Bangiales, Rhodophyta)

The optimal conditions for growth of Porphyra dioica gametophytes were investigated in the laboratory, focusing on bioremediation potential. Porphyra dioica is one of the most common Porphyra species along the northern coast of Portugal and can be found year-round. The influence of stocking density and photon flux density (PFD) on the growth, production and nutrient removal was tested. Maximum growth rates, up to 33% per day, were recorded with 0.1 g fw l^− 1 at 150 and 250 μmol photons m^− 2 s^− 1. Growth rate decreased significantly with increasing stocking density. Productivity (g fw week^− 1) had an inverse trend, with more production at the higher stocking densities. At 150 μmol m^− 2 s^− 1 and with 1.5 g fw l^− 1, 1.4 g fw week^− 1 were produced. At this PFD, there was no significant difference in production between 0.6 to 1.5 g fw l^− 1. Nitrogen (N) content of the seaweeds decreased with increasing stocking densities and PFDs. The maximum N removal was recorded at 150 μmol m^− 2 s^− 1, with 1.5 g fw l^− 1 stocking density (1.67 mg N day^− 1). However, the N removed by thalli at 50 μmol photons m^− 2 s^− 1 was statistically equal to that at 150 and 250 μmol photons m^− 2 s^− 1, at a stocking density of 1.0 g fw l^− 1. The influence of temperature and photoperiod on growth and reproduction was also assessed. Growth rates of P. dioica were significantly affected by temperature and photoperiod. In this experiment (with 0.3 g fw l^− 1 stocking density), the highest growth rate, 27.5% fw day^− 1, was recorded at 15 °C and 16 : 8¯, L : D¯. Male thalli started to release spermatia 21 days after the beginning of the experiment, in temperatures from 10 to 20 °C and with 10, 12 and 16 h of day length. Unfertilized female-like thalli were observed at 10 to 20 °C, under all photoperiods tested. Growth of these thalli declined after 4 weeks. By then, formation of young bladelets in the basal portion of these thalli was observed. After 7 weeks all biomass produced was solely due to these vegetatively propagated young thalli, growing 22.4% to 26.1% day^− 1. The results of this study showed that P. dioica appears to be a candidate as a nutrient scrubber in integrated aquaculture systems.     

The effects of light and temperature on the photosynthesis of the Asparagopsis armata tetrasporophyte (Falkenbergia rufolanosa), cultivated in tanks

The integrated aquaculture of the tetrasporophyte of Asparagopsis armata Harvey (Falkenbergia rufolanosa) using fish farm effluents may be viable due to the species high capacity of removing nutrients and its content of halogenated organic compounds with applications on the pharmaceutical and chemical industries. In order to optimize the integrated aquaculture of F. rufolanosa, we followed the daily variation of the potential quantum yield (F_v/F_m) of PSII on plants cultivated at different biomass densities and different total ammonia nitrogen (TAN) fluxes to check if they are photoinhibited at any time of the day. Moreover, the photoinhibition under continuous exposure to highly saturating irradiance and its potential for subsequent recovery in the shade was assessed. The potential for year round cultivation was evaluated by measuring rates of O₂ evolution of plants acclimated at temperatures ranging from 15 to 29 °C, the temperature range of a fish farm effluent in southern Portugal where an integrated aquaculture system of F. rufolanosa was constructed.Photoinhibition does not seem to be a major constrain for the integrated aquaculture of F. rufolanosa. Only when cultivated at a very low density of 1.5 g fresh weight (FW) l^− 1 that there was a midday decrease in maximal quantum yield (F_v/F_m). At densities higher than 4 g FW l^− 1, no photoinhibition was observed. When exposed to full solar irradiance for 1 h, F. rufolanosa showed a 33% decrease in F_v/F_m, recovering to 86% of the initial value after 2 h in the shade. A midday decline of the F. rufolanosa F_v/F_m was also observed under the lowest TAN flux tested (∼6 μM h^− 1), suggesting that this fast and easy measurement of fluorescence may be used as a convenient diagnostic tool to detect nutrient-starved unbalance conditions of the cultures. Maximum net photosynthesis peaked at 15 °C with 9.7 mg O₂ g dry weight (DW)^− 1 h^− 1 and remained high until 24 °C. At 29 °C, the net oxygen production was significantly reduced due to a dramatic increase of respiration, suggesting this to be the species' lethal temperature threshold.Results indicate that F. rufolanosa has a considerable photosynthetic plasticity and confirm it as a good candidate for integrated aquaculture at temperatures up to 24° C and cultivation densities of at least 5 g FW l^− 1. When cultivated at these densities, light does not penetrate below the first few centimetres of the surface zone. Plants circulate within the tanks, spending around 10% of the time in the first few centimetres where they are able to use efficiently the saturating light levels without damaging their photosynthetic apparatus.     

Successful culture of larvae of Litopenaeus vannamei fed a microbound formulated diet exclusively from either stage PZ2 or M1 to PL1

In three separate experiments, harpaticoid copepods Tisbe monozota (alive and dead) and a microparticulate microbound diet were evaluated as alternatives to live Artemia nauplii as food, beginning at either stage PZ2 or M1, in the larval culture of Litopenaeus vannamei. Larvae were cultured in 2 L round bottom flasks at a density of 150 L^− 1 (Experiment 1) and 100 L^− 1 ( 3.2 and 3.3) at 28 °C, 35‰ salinity and 12:12 LD photoperiod, and fed 4×/day^- 1. Larvae were initially fed a mixture of phytoplankton to stages PZ2 or M1 and then fed either live Artemia, live or dead copepods, or a microparticulate microbound diet. The experiments were terminated and all larvae were harvested when more than 80% of larvae had molted to postlarvae 1 (PL1) within any flask representing any of the treatments. The comparative value of the different diets and feeding regimes was determined by mean survival, mean dry weight and total length of individual larva, and percentage of surviving larvae that were PL1. Trypsin activity of samples of larvae from each treatment was also determined. The microparticulate microbound diet effectively served as a complete substitute for Artemia nauplii when fed beginning at stage M1. When fed at the beginning of the PZ2 stage, survival was comparable to that of larvae fed Artemia, but mean dry weight, mean total length, and percent of surviving larvae that were PL1 generally were significantly less. Responses to the feeding of copepods, whether fed dead or live, as a substitute were generally significantly less than those of larvae fed either the Artemia nauplii or the microparticulate diet. Values of trypsin activity (10^− 5 IU/μg^- 1 dry weight) corresponded to the relative proportions of the different larval stages within a treatment, with higher activity being characteristic of early stages. Previously demonstrated successful results with another species of crustacean suggest that the microparticulate microbound diet has characteristics that should be effective in the culture of the carnivorous stages of other crustacean and fish larvae that are currently fed live Artemia nauplii.     

First feeding of Octopus vulgaris Cuvier, 1797 paralarvae using Artemia: Effect of prey size, prey density and feeding frequency

Different assays related to the first feeding of Octopus vulgaris Cuvier, 1797 are compiled in this paper. They include: age at initial feeding age, prey size selection and optimal density, attack timing after feeding, and effect of dose number on the number of captures. Prey capture and ingestion processes were also analysed. Food supplied was cultured Artemia sp. Each assay lasted 15 min.Although paralarvae already start to feed on the hatching day (day 0), it is during day 2 when a greater number of attacks is recorded (81.7 ± 14.7% paralarvae attack). They mainly prefer (significance level α = 0.05) large Artemia, 1.4 ± 0.4 mm (77.0 ± 5.6% of the total attacks) than small Artemia, 0.8 ± 0.1 mm (23.0 ± 5.6%). There is also a slight predilection for the lowest Artemia concentration (33.3 ±12.6% paralarvae attack in a 0.1 Artemia ml^− 1 density, opposite 16.7 ± 7.6 and 18.3 ± 7.6% in densities of 0.5 and 1 Artemia ml^− 1 respectively). The greatest predatory activity is recorded during the first 5 min after food is supplied (72.2 ± 25.5%). An increase in the predatory activity was also observed when food was distributed in several doses instead of a single dose (75.0 ± 10.0% and 46.7 ± 17.6% respectively). It was proved for the first time that paralarvae completely ingest their preys (including their exoskeletons), in this case Artemia. Time needed for their total ingestion ranges between 4 and 10 min.     

Nitrogen and phosphorous budgets during a farming cycle of the Manila clam Ruditapes philippinarum: An in situ experiment

In the Sacca di Goro lagoon a farming cycle of the Manila clam (Ruditapes philippinarum) was simulated seeding young molluscs in an unexploited sandy spot. The experimental area (2100 m²) consisted of three sectors: a control (C), almost devoid of clams (∼ 1600 m^− 2, ∼ 30 ind m^− 2), a low (L) density area (400 m², ∼ 300 ind m^− 2) and a high (H) density zone (∼ 110 m^− 2, ∼ 800 ind m^− 2). Water chemistry, external freshwater nutrient loads, molluscs filtration rates, biomass, elemental composition and nutrient recycling were analysed.Clam filtration rates and light and dark fluxes of nutrients were measured with intact core incubations. Three replicate cores (i.d. 20 cm) were collected from C, L and H in April, one month after the seeding, June, August and October 2003. External loads were calculated multiplying dissolved and particulate nutrients concentration by freshwater flow from the main lagoon tributaries. Direct excretion, filtration activity of clams and particulate matter deposition resulted in significantly higher ammonium (NH₄⁺) and soluble reactive phosphorus (SRP) effluxes to the water column at L and H. For the entire farming cycle, particulate nitrogen (PN) uptake by clams from the water column was 1.7 (C), 9.1 (L) and 16.3 (H) mol m^− 2, whilst total dissolved nitrogen (TDN) fluxes were − 0.3 (C), 1.6 (L) and 6.9 (H) mol m^− 2. Particulate phosphorus (PP) uptake from the water column was 0.1 (C), 0.6 (L) and 1.0 (H) mol m^− 2, whilst total dissolved phosphorus (TDP) efflux was 0.2 (C), 0.5 (L) and 0.8 (H) mol m^− 2. At the end of the farming cycle, harvested N as mollusc flesh was negligible for C, 0.4 mol m^− 2 for L and 1.8 mol m^− 2 for H. Harvested P as mollusc flesh was negligible for C, 0.02 mol m^− 2 for L and 0.04 mol m^− 2 for H. Farmed areas seem to have a great potential for fast coupling between sedimentation (filter feeder mediated biodeposition) and benthic recycling. At the lagoon level, mollusc farming probably attenuates the export of particulate matter to the open sea. Our results show that a minor fraction of biodeposited N (∼ 6%) and P (∼ 3%) was exported as a commercial product at the end of the farming cycle, whilst a larger fraction was incorporated in the sediments or recycled as dissolved inorganic or organic forms.     

Oxytetracycline (OTC) accumulation and elimination in hemolymph, muscle and hepatopancreas of white shrimp Litopenaeus vannamei following an OTC-feed therapeutic treatment

The white shrimp, Litopenaeus vannamei, has become a very important species for the development of shrimp aquaculture in Northwest Mexico. However, viral and bacterial diseases are considered a major threat to the development of this industry. In the present study a trial was conducted to evaluate the tissue distribution, maximum concentration, and elimination of the widely used antibiotic oxytetracycline (OTC) in L. vannamei using indoor tanks under laboratory-controlled conditions. OTC was given to shrimp simulating a therapeutic treatment through medicated feed for 14 days followed by a period of feeding without antibiotic for another 14 days to evaluate the elimination pattern. Samples of hemolymph, muscle, and hepatopancreas were taken from medicated animals every two days for 28 days. All tissues were removed and frozen immediately in liquid nitrogen. OTC levels were analyzed by High Performance Liquid Chromatography (HPLC). Results showed an important OTC increase during consumption of medicated feed in all examined tissues. OTC maximum concentrations were 33.54 ± 11.19, 194.37 ± 16.11, and 18.79 ± 5.87 µg g^− 1 for muscle, hepatopancreas and hemolymph, respectively. Although the highest OTC level was found in the hepatopancreas, it required only two days after the start of dosing to reach this value, whereas the maximum OTC for muscle and hemolymph was detected after eight days of dosing. Ten days after the cessation of medicated feeding, the drug content in the shrimp tail muscle was under the detectable limit for the method (0.01 µg g^− 1 of OTC).     

Experimental integrated aquaculture of fish and red seaweeds in Northern Portugal

Three potentially valuable red seaweeds, Chondrus crispus Stackhouse, Gracilaria bursa pastoris (S.G. Gmelin) P.C. Silva and Palmaria palmata (L.) O. Kuntze, collected in northern Portugal, were cultivated using the nutrient-rich effluents from a local turbot (Scophthalmus maximus Linnaeus) and sea bass (Dicentrarchus labrax Linnaeus) farm. The algae were cultivated in a two level cascade system. Several arrangements of the cascade system, stocking densities (3, 5, 7 and 8 kg m^− 2) and water fluxes (140 and 325 l h^− 1) were tested to optimize biomass yield and nitrogen uptake rate and efficiency. The yield and the total ammonium nitrogen (TAN) uptake of the three species were highly seasonal. Palmaria could not survive culture conditions during the summer when water temperature was above 21 °C. In the spring, Palmaria had an average yield of 40.2 (± 12.80) g DW m^− 2 day^− 1 and a nitrogen uptake efficiency (NUE) of 41.0% (± 17.26%). NUE expresses, in percentage, the average reduction in TAN concentration between the inflows and the outflows of the tanks. Chondrus performed better in summer with an average yield of 37.0 (± 11.10) g DW m^− 2 day^− 1 and removing 41.3% (± 17.32%) of nitrogen. Gracilaria grew year round, but also performed better during spring/summer, producing an average of 29.1 (± 2.90) g DW m^− 2 day^− 1, and only 7.3 (± 5.08) g DW m^− 2 day^− 1 during autumn. Yield of C. crispus did not differ significantly when grown at two different stocking densities (5 kg m^− 2 and 8 kg m^− 2). On the other hand, Gracilaria had significantly higher yields at 5 than at 7 kg m^− 2. Better NUE, on average 76.7% (± 22.13%), was also obtained with 5 kg m^− 2 stocking density and only 63.8% (± 24.62%) with 7 kg m^− 2. The yield of Gracilaria increased significantly with the increase of water flux from 140 to 325 l h^− 1 and more nitrogen was removed from the water. However, NUE decreased from 48.4% to 33.4% at 140 and 325 l h^− 1, respectively. Biofiltration was highly improved by a cascade system with a NUE as high as 83.5%.     

Routes of water loss in South African abalone (Haliotis midae) during aerial exposure

South African abalone, Haliotis midae, were exposed to air at 12 °C for 36 h to simulate the extent and rate mass loss experienced by animals during long distance live exports. Animals lost 15.1 ± 0.94% of their mass during the 36 h air exposure, an approximation of the highest mass losses sustained by industry.The total mass loss was attributed to water loss, as the contribution of dry mass to the total mass remained constant under all conditions. Water content decreased from 64.8% of the body mass (M_b) under control conditions to 58.8% M_b after 36 h in air. In real terms, however, animals had lost 22% of the body water pool.Abalone exhibited a typically high water turnover rate when in water (125 μL g^− 1 h^− 1), which decreased markedly during air exposure (2.2 μL g^− 1 h^− 1). Haemolymph volume decreased from 43% M_b in water to 14% M_b in air. The concomitant decrease in haemolymph pressure probably limited the first step in urine formation (ultra-filtration through the pericardium). Thus we observed that while urine flow represented about 26% of the total water loss when the animals were in water, urine flow ceased during air exposure.The decrease in haemolymph volume in air represents a redistribution of water to the tissues and not a bulk loss of haemolymph. This is supported by the concentration of haemolymph ions by a factor of 1.2 during aerial exposure, which was predicted based on the 22% decrease in water content. Under the same conditions, evaporation from water containers with similar surface to volume dimensions as abalone, accounted for only an 8.25% mass loss. As all other water loss routes were accounted for, we measured pedal mucus production rates of abalone in water and air. During 36 h aerial exposure, the pedal mucus production represented a loss of 6.8% M_b. We conclude that water loss during 36 h air exposure is attributable to evaporation (8.25% M_b) and pedal mucus production (6.8% M_b). This paves the way for directed research into mitigating water loss during the live export process.     

Productivity improvement of Lysmata seticaudata (Risso, 1816) larval rearing protocol through modelling

The Monaco shrimp Lysmata seticaudata (Risso, 1816) is a marine ornamental species whose ecology and biology, as well as its larval culture has previously been addressed. The objective of the study was to predict and improve productivity of this species rearing protocol through modelling. The models developed intend to help aquaculturists to maximize survival to postlarva, decrease larval duration and increase synchronism of metamorphosis and newly metamorphosed postlarvae size by manipulating temperature, diet, first feeding period and stocking density.The models developed allow us to conclude that the L. seticaudata rearing protocol productivity can be improved by raising larvae at a density of 40 larvae L^− 1 and fed newly hatched Artemia nauplii since hatching to zoea V, and with Algamac 2000™ enriched Artemia metanauplii from zoea V to metamorphosis to postlarvae.By providing more productive protocols to aquaculturists, destructive practices and wild collection may be reduced.     

Effects of dietary vitamin C on survival, growth, and immunity of large yellow croaker, Pseudosciaena crocea

An 8 weeks growth experiment was conducted to determine the effects of dietary vitamin C on the survival, growth, tissue ascorbic acid contents and immunity of large yellow croaker (Pseudosciaena crocea) with initial weight of 17.82 ± 0.68 g. Seven practical diets were formulated to contain 0.1, 12.2, 23.8, 47.6, 89.7, 188.5 and 489.0 mg ascorbic acid equivalent kg^− 1 diet, supplied as l-ascorbyl-2-polyphosphate (LAPP). Each diet was fed to triplicate groups of fish in seawater floating cages (1.5 × 1.5 × 2.0 m), and each cage was stocked with 120 fish. Fish were fed twice daily (05:00 and 17:00) to apparent satiation for 8 weeks. The water temperature fluctuated from 19.5 to 25.5 °C, the salinity from 25 to 28‰ and dissolved oxygen content was approximately 7 mg l^− 1 during the experimental period. Results showed that the specific growth rate (SGR) (from 1.80 to 1.96% d^− 1) had an increasing trend with the increase of dietary vitamin C, but no significant difference was observed among dietary treatments. No gross deficiency signs were observed in any of the experimental fish. Survival rate, however, significantly increased with increasing dietary vitamin C (P < 0.05). The vitamin C contents in liver and muscle correlated positively with the vitamin C in diets. The vitamin C requirement was estimated to be 28.2 mg kg^− 1 based on survival, and 87.0 mg kg^− 1 on liver content of vitamin C. The activities of serum lysozyme and alternative complement pathway (ACP), phagocytosis percentage (PP) and respiratory burst activity of head kidney significantly increased with increasing dietary vitamin C. The challenge experiment with Vibrio harveyi showed that fish fed the diets with supplementation of vitamin C had significantly lower cumulative mortality compared to the control group (66.7%), and the cumulative mortality (16.7%) in fish with 489.0 mg kg^− 1 ascorbic acid was significantly lower than that (41.7%) in fish with 23.8 mg kg^− 1 ascorbic acid. These results suggested that vitamin C significantly influenced the immune response and disease resistance of large yellow croaker.     

Detection of V. harveyi in shrimp postlarvae and hatchery tank water by the Most Probable Number technique with PCR

V. harveyi is the cause of serious disease in the shrimp industry in Thailand during cultivation. In this study, the gyrB gene of V. harveyi NICA, isolated from shrimp in Thailand, was sequenced. A pair of specific primers (A2B3) was designed that allowed amplification of a 363 bp gene fragment of V. harveyi. No cross reaction was detected in 17 other Vibrio species tested except for V. carchariae which is a synonym for V. harveyi. The possibility of using A2B3 for confirmation and enumeration of V. harveyi by PCR was demonstrated. Of 40 possible V. harveyi strains isolated from seafood on the basis of their growth on TCBS plates and biochemical reactions, 36 gave a reaction with the specific primers. The primers could detect V. harveyi at a level of as few as 15 cells/ml. The Most Probable Number (MPN) technique was applied to enumerate V. harveyi. We have demonstrated that when PCR was applied directly to the enrichment broth of shrimp artificially inoculated with V. harveyi, the MPN value was no different from the MPN value obtained using the standard technique with selective agar. This technique was employed to enumerate V. harveyi in postlarvae and hatchery tank water. V. harveyi were detected in 18 out of 21 postlarval samples and in 14 out of 21 tank water samples. The numbers of V. harveyi detected in postlarvae and water were 150-1.1 × 10⁸/g postlarvae and 7-4.6 × 10⁴/ml of water samples, respectively. Screening of postlarvae to reduce the high risk of V. harveyi contamination in cultivation ponds is suggested as a measure to prevent the catastrophic losses caused by V. harveyi disease.     

Vegetative propagation of the carrageenophytic red alga Gigartina skottsbergii Setchell et Gardner: Indoor and field experiments

The carrageenophytic red alga Gigartina skottsbergii presents several biological constraints for its cultivation such as restricted temporal availability and high spore mortalities that affect the development of its mass cultivation. For this reason, research to develop alternative propagation methods has been undertaken. Previous laboratory studies demonstrated that manipulating temperature, light and nutrients could enhance healing and regeneration of this seaweed. In this study nursery and field experiments were undertaken to establish the possibility to regrow G. skottsbergii in conditions similar to those applied in mass cultivation practices. Frond fragments and rhizoids were tested as alternative ways to obtain new plants. Our results indicate that regeneration occurs in the field, and can be managed in nursery conditions. The addition of a nitrogen source enhances the growth from 0.6 to 1.0% d^− 1 of the healed fragments and the use of a photon flux density above 50 μmol m^− 2 s^− 1 increases the growth rate but decrease the survival of the explants. Explants can be transferred to field conditions and grow at similar rates to those registered in the nursery (0.5% d^− 1). It is also possible to propagate rhizoids of G. skottsbergii and it seems that the attachment of a frond portion to the substratum, enhanced its survival and regeneration capacity. Finally, this study demonstrated that rhizoids attached to rocks can regenerate complete plants in nature, a feature which could be a useful for developing a sustainable harvesting methodology.     

Effect of temperature on growth and energy budget of juvenile cobia (Rachycentron canadum)

Growth and energy budget of juvenile cobia (initial body weight ∼ 22 g) at various temperatures (23, 27, 31 and 35 °C) were investigated in this study. Maximal ration level (RL_max, %/day) increased as temperature (T, °C) increased from 23 °C to 31 °C but decreased at 35 °C, described as a quadratic equation: RL_max = −0.023T² + 1.495T − 17.52. Faecal production (f, mg g^− 1 day^− 1) increased with increased temperature (T, °C), described as a power function: lnf = 0.738lnT − 0.806. As temperature increased, feed absorption efficiency in dry weight (FAE_d, %), protein (FAE_p, %) and energy (FAE_e, %) all increased first and then decreased, but the variation of feed absorption efficiency was small, with ranges of 89.59-91.08%, 92.91-94.71%, 93.92-95.32%, respectively. Specific growth rate in wet weight (SGR_w, %/day), dry weight (SGR_d, %/day), protein (SGR_p, %/day) and energy (SGR_e, %/day) showed a domed curve relative to temperature (T, °C), described as quadratic equations: SGR_w = − 0.068T² + 3.878T − 50.53, SGR_d = − 0.079T² + 4.536T −59.64, SGR_p = − 0.084T² + 4.783T − 63.08 and SGR_e = − 0.082T² + 4.654T − 60.99, and SGR_w, SGR_d, SGR_p and SGR_e maximized at 28.5 °C, 28.6 °C, 28.4 °C, 28.5 °C, respectively, as calculated from the regression equations. The relationships between feed conversion efficiency in wet weight (FCE_w, %), dry weight (FCE_d, %), protein (FCE_p, %), energy (FCE_e, %) and temperature (T, °C) also took on a domed curve described as quadratic equations: FCE_w = − 0.726T² + 39.71T − 473.8, FCE_d = − 0.276T² + 15.31T − 190.6, FCE_p = − 0.397T² + 22.05T − 277.9 and FCE_e = − 0.350T² + 19.39T − 239.9, and FCE_w, FCE_d, FCE_p and FCE_e maximized at 27.4 °C, 27.8 °C, 27.7 °C and 27.7 °C, respectively, as calculated from the regression equations. Energy budget of juvenile cobia fed satiation was 100C = 5F + 67(U + R) + 28G at water temperature 27 °C and 100C = 5F + 70(U + R) + 25G at water temperature 31 °C, where C is food energy, F is faeces energy, (U + R) is excretion energy and metabolism energy, and G is growth energy.     

Towards development of large-scale hatchery cultivation of larvae and spat of the pearl oyster Pinctada mazatlanica in Mexico

The results of a series of pilot-scale runs with P. mazatlanica larvae from 2004 through 2006 are reported. Preliminary runs in 2004 and 2005 used broodstock collected in summer, when massive spawning of wild populations naturally occurs. However, results of larval development were very poor and failed to produce spat in both years. In 2006, ripe broodstock were still collected in summer, but also in the spring time, based on the hypothesis that the gonads in this season were in better reproductive condition that in summer. Three larval runs were conducted in 2006: two in spring and one in summer. Larvae growth and survival greatly increased in both spring runs, ending with two successful productions of spat (∼ 20 × 10³ and ∼ 100 × 10³ juveniles). The summer larval run in 2006 failed again to produce spat. Additionally, the first run of April 2006 refers to an experiment that evaluated two different larval culture conditions: constant temperature (27 °C) and low stocking density (3-4 larvae ml^− 1) versus variable temperature (24-28 °C) and high stocking density (8-9 larvae ml^− 1). The first trial significantly increased larval survival and growth, which in turn resulted in greater numbers of settled spat, in comparison of the second trial, where survival, growth, and settlement of spat were significantly lower. Also in 2006, the quality of seawater used at the hatchery was evaluated with microbiological and chemical tests. The implication of these tests, together with results from all experiments are analyzed and discussed in terms of the potential development of large-scale hatchery cultivation of P. mazatlanica larvae in Mexico.     

An insulin-like system involved in the control of Pacific oyster Crassostrea gigas reproduction: hrIGF-1 effect on germinal cell proliferation and maturation associated with expression of an homologous insulin receptor-related receptor

The putative involvement of insulin-like peptides in the control of the reproduction of the Pacific oyster Crassostrea gigas was investigated using different approaches. In conjunction with a monthly histological analysis of the oyster reproductive cycle, in vitro biological effects of the human recombinant IGF-1 (hrIGF-1) on dissociated germinal cells were mesured over 1 year using [³H]-thymidine and [¹⁴C]-amino acid mixture as tracers for DNA and protein synthesis. DNA synthesis was stimulated by hrIGF-1 in November (114 ± 11% for 10^− 7M), December (46 ± 6% for 10^− 7 M) and January, which was identified as the highest gonial mitosis period. A clear dose-effect was observed in January with a maximum activation of 68 ± 7% for 10^− 12 M. Germinal cell protein synthesis was also stimulated in March (20 ± 1% for 10^− 10 M), April (41 ± 5% for 10^− 13 M), May (25 ± 4% for 10^− 13 M), and by almost all of hrIGF-1 doses in June (21.5 ± 2% for 10^− 13 M) and July (34 ± 1% for 10^− 13 M). This suggests the involvement of insulin-like substances in gonadal tubule rebuilding (December), as well as in the development of germinal cells (March, April), and in the summer maturation of gametes (May, June, July). These insulin-like effects conform with the expression pattern of the recently identified C. gigas insulin receptor-related receptor (CIR): It appeared highly expressed in the gonadal area during gonial mitosis phase, but also in maturating oocytes, suggesting the involvement of an insulin-like system in gonial proliferation and maturation. Moreover, CIR showed differential expression during embryogenesis and larval developmental stages. The expression of maternal CIR during the embryonic and early larval development, followed by the increasing zygotic CIR expression from D larvae to 11-day-old veliger larvae, then a decrease until metamorphosis, also suggest that insulin-like peptide is involved in organogenesis.