Comparison of diagnosis techniques for the protozoan parasite Bonamia ostreae in flat oyster Ostrea edulis

In recent years, the cultured production of the flat oyster Ostrea edulis has suffered a dramatic decrease in Europe partially attributed to the protozoan parasite Bonamia ostreae, the causative agent of the Bonamiosis. In this paper the results of a PCR assay for the diagnosis of B. ostreae were compared with those obtained using two classical methods of diagnosis recommended by the OIE (Office International des Epizooties) and the European Union, histology and cytology. The same samples were analyzed by two different laboratories, showing that the results obtained with the PCR method have high sensitivity and good correlation between laboratories. This method is cheaper and faster than histopathology and cytology, with no need of specifically trained personnel to perform the diagnoses. It is appropriate for fast screening of stocks of great numbers of oysters.     

Distribution of a fish pathogen Listonella anguillarum in the Japanese flounder Paralichthys olivaceus hatchery

The present study was undertaken to investigate the distribution of Listonella anguillarum in a Japanese flounder (Paralichthys olivaceus) hatchery. A total of 2704 isolates were obtained from the developing fish, live diets and artificial feeds of Japanese flounder and their rearing water, 439 of which were identified as L. anguillarum by the combining incubation on thiosulfate-citrate-bile salt-sucrose (TCBS) agar at 35 °C overnight with polymerase chain reaction (PCR) detection for the VAH1 hemolysin gene. L. anguillarum was detected in all seven rotifer samples, with densities of 2.5 × 10³ to 4.6 × 10⁶ colony forming units (CFU) g^− 1. Both the analyzed samples of Nannochloropsis oculata contained this bacterium at densities of 1.6 × 10⁴ to 1.4 × 10⁵ CFU g^− 1. L. anguillarum was detected in only one of four samples of Artemia nauplii with a density of 4.8 × 10⁵ CFU g^− 1 (35%) and it was not detected in the two analyzed artificial feed samples. L. anguillarum was detected in 11 of 18 specimens of larval and juvenile Japanese flounder at densities of 5.0 × 10¹ to 7.4 × 10⁵ CFU g^− 1, while it was not detected in the two analyzed egg specimens of Japanese flounder. These results indicate that L. anguillarum associated with the developing Japanese flounder is likely derived from rearing water and live diets such as rotifers. Further, it is strongly suggested that L. anguillarum is a transient bacterium of the intestinal microflora for the Japanese flounder but is a permanently indigenous one for the Japanese flounder hatcheries.     

Selective particle feeding by the Chilean oyster, Ostrea chilensis; implications for nursery culture and broodstock conditioning

Hatchery broodstock conditioning and nursery culture of the Chilean flat oyster Ostrea chilensis have been hampered by the poor performance of oysters fed typical microalgal hatchery diets. To determine the feeding capabilities of this species the selective removal and consumption of natural planktonic assemblages and artificial inert particles (polystyrene beads) by juvenile and adult oysters were examined experimentally. The arrangement of the eulaterofrontal cirri of the ctenidia was also examined to infer their potential efficiency of particle selection for feeding. Polystyrene beads of 45 and 15 μm in diameter had high rates of removal from suspensions by both juvenile (45 μm = 70%, 15 μm = 73%) and adult (45 μm = 88%, 15 μm = 87%) oysters. In contrast, beads of 6 μm diameter had lower rates of removal (adults = 68%, juveniles = 53%), while 1 μm beads were not removed at all. Both adult and juvenile oysters feeding upon natural plankton assemblages removed only microphytoplankton (20-75 μm in size) despite the presence of nanophytoplankton (2-20 μm), picophytoplankton (< 2 μm), cyanobacterium Synechoccocus spp. (< 2 μm), and bacterial cells (< 75 μm) in the experimental suspensions. Eulaterofrontal cirri of both juvenile (15.2 μm ± 0.9 SE) and adult oysters (18.9 μm ± 0.3 SE) are the shortest that have been reported for any ostreid species which helps to explain the inability of this species to retain small food particles. The clearance rates for oysters feeding on microphytoplankton only were 1.49 (± 0.05 SE) and 7.1 (± 1.2 SE) l h^− 1g^− 1 for juveniles and adults respectively. These values are much higher than previously reported for this species being fed smaller sized cultured microalgae. Our results strongly suggest that the difficulties in the nursery and broodstock hatchery culture of this oyster may be due to inappropriate phytoplankton diets. We recommend the provision of cultured microalgae of 20-75 μm in diameter for improving the performance of hatchery maintained juvenile and adult O. chilensis.     

Resistance of Ostrea edulis to Bonamia ostreae infection

Preliminary studies showed that oysters from a population of Ostrea edulis infected with Bonamia ostreae since at least 1963 have a substantial degree of resistance to the infection in comparison with oysters from a population with no prior exposure history to the disease. Over a 46-week period and under identical environmental conditions, 99% of non-resistant oysters exposed to the disease died while only 26% of a population of resistant oysters died. The surviving resistant oysters had lesions which indicated a chronic infection and possible degradation of the parasite within the tissues. The resistant oysters may offer a management option for the reestablishment of Ostrea edulis culture in disease enzootic areas where the parasite has virtually eliminated non-resistant populations.     

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.     

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.     

Evaluation of three seaweeds Gracilaria bursa-pastoris, Ulva rigida and Gracilaria cornea as dietary ingredients in European sea bass (Dicentrarchus labrax) juveniles

The aim of this study was to evaluate the inclusion of three seaweeds Gracilaria bursa-pastoris (GP), Ulva rigida (UR) and Gracilaria cornea (GC) as dietary ingredients on the performance, nutrient utilisation and body composition of European sea bass juveniles. Six experimental diets were formulated to replace 5% (GP-5, UR-5, and GC-5 Diets) and 10% (GP-10, UR-10 and GC-10 Diets) fish protein hydrolysate (CPSP) by each of the three seaweeds. A control diet was used, without inclusion of any seaweed. Diets were fed to duplicate groups of 25 European sea bass (Dicentrarchus labrax) juveniles (IBW = 4.7 g) for 10 weeks. Growth performance was only significantly reduced (P < 0.05) in fish fed the GC-10 diet, whereas the feed conversion ratio increased significantly in those fish. The apparent digestibility coefficients of dry matter and lipid were significantly lower in fish fed diet GC-10 relative to those fed the control diet. Carcass composition was similar among treatments, although fish fed GC-10 exhibited significantly higher ash content.The results obtained in this study suggest that the inclusion of G. bursa-pastoris (GP) and U. rigida (UR), up to 10%, can be considered as very interesting ingredients in diets for sea bass juveniles, as no negative consequences on growth performance, nutrient utilization or body composition were observed. On the other hand, the inclusion of G. cornea (GC) should be limited to 5% of the diet.     

Recruitment and effects of Discocotyle sagittata (Monogenea) infection on farmed trout

Farmed rainbow trout Oncorhynchus mykiss and brown trout Salmo trutta were monitored over 3 years for infection with the blood-feeding gill fluke Discocotyle sagittata. Parasite transmission is seasonal: new infections take place during summer/autumn, and transmission is generally negligible during winter/spring. There are 2 sources of infection for naïve fish-of-the-year: limited invasion when fish are in the raceways by riverborne larvae originating external to the farm; and internally, within the farm, when 0+ fish are transferred to ponds previously occupied by older cohorts of infected fish. Thereafter, infection levels continue to increase in rainbow trout primarily through transmission within the farm. Prevalence rose to 100% in 1+ fish by the end of their second summer. In O. mykiss, mean abundance reached 194 worms/host for 1+ fish (up to 489 worms/host) and 160 worms/host for 2+ fish. By contrast, in S. trutta, parasite prevalence never exceeded 85% and, after the first year's invasions, infection levels decreased over time: in 1+ and 2+ brown trout, parasite mean abundance was < 4 (maximum 15) worms/host. We present evidence of the detrimental effects of D. sagittata on the host: high burdens are associated with pale gills, decreased body condition and host mortality. Parasite burdens become overdispersed during the warmer part of the year, as prevalence and mean abundance increase. However, the degree of parasite overdispersion decreases over winter; we cannot distinguish whether decreased aggregation is due to parasite losses from infected fish (including immune-mediated parasite mortality) or parasite-induced host mortality.     

Suitability of the copepod, Acartia clausi as a live feed for Seabass larvae (Lates calcarifer Bloch): Compared to traditional live-food organisms with special emphasis on the nutritional value

Though artificial propagation of Asian seabass Lates calcarifer (Bloch) in captivity through induced breeding techniques is standardized under Indian conditions, larval and nursery rearing techniques including suitable nursery feeds have to be standardized to obtain better survival and growth. Feeding experiments in triplicate were conducted to evaluate the suitability of the marine copepod Acartia clausi as live prey for fourteen day-old seabass larvae (6.53 ± 0.06 mm; 8.58 ± 0.33 mg) and compared with the traditional live prey, rotifers and Artemia nauplii. While A. clausi and rotifers were mass produced using algae Isochrysis galbana, Chaetoceros affinis and Chlorella marina, Artemia nauplii were produced using cysts. Nutritional quality of cultured copepods was evaluated based on the proximate composition, amino acid and fatty acid composition, and compared with that of rotifers and Artemia nauplii. Proximate composition varied significantly (P < 0.05) among the different live feeds. A. clausi showed higher protein (63.12%) and lipid (16.65%) content than Artemia nauplii and rotifers. Total essential amino acids content was 2% lower in A. clausi compared to that in Artemia nauplii. Fatty acid profiles of the live feed organisms showed that A. clausi is a rich source of n − 3 fatty acids. The total n − 3 fatty acid content of A. clausi was 33.94%. Length, weight overall weight gain and survivorship were significantly (P < 0.05) different among the dietary treatments, and weight gain was comparatively higher in A. clausi fed larvae. Survival of seabass larvae fed A. clausi was obtained highest as 58.13% against the lower values of 39.93% and 41.62% in larvae fed rotifer and Artemia nauplii respectively. Final carcass composition of the larvae of L. calcarifer fed different live-food organisms showed significant differences (P < 0.05) among the dietary treatments. The fatty acid composition of the dietary treatments was reflected to a certain extent in the fatty acid composition of the seabass larvae. The present investigation revealed the nutritional value of calanoid copepod and thus underlining its usefulness as a suitable live-food organism for rearing larvae of the commercially valuable Asian seabass.     

Prey ingestion and live food selectivity of marble goby (Oxyeleotris marmorata) using rice field prawn (Macrobrachium lanchesteri) as prey

Marble goby (Oxyeleotris marmorata), a carnivorous fish native to freshwater in Asia-Pacific region, is a high-valued species in many Asian countries. The present study consisting of three experiments was conducted to determine the appropriate density, size and ingestion time of marble goby fingerlings on rice field prawn (Macrobrachium lanchesteri) as prey. Results showed that the ingestion rate of marble goby fingerlings (0.26-1.43 g size) on prawns (9-12 mm in length) had an asymptotic pattern peaked at a predator to prey ratio of 1:20. Marble goby fingerlings did catch prawns during both daytime (07:00-17:00 h) and nighttime (17:00-07:00 h), but the ingestion peaked during nighttime period. The hourly ingestion rate during nighttime was significantly higher than that during daytime for the smaller-size marble goby fingerlings of 0.26-0.87 g, while there was no significant difference in the hourly ingestion between nighttime and daytime for the larger-size marble goby fingerlings of 1.43 g. The daily ingestion of marble goby fingerlings on prawns (mg consumed prawns/goby/day) increased significantly with increasing average weights (g) of marble goby fingerlings (Y = 27.6 + 47.8X, n = 30, r² = 0.86, p < 0.05). Results also showed that the preferred prawn size of marble goby fingerlings of average weights of 0.26-4.18 g was small (9-12 mm) with positive electivity index of 0.19-0.33, and the shift from small- to medium-size (12-14 mm) prawns was slow. The present study has demonstrated the feasibility of using rice field prawns as live foods to nurse marble goby fingerlings.     

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.     

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.     

Development and evaluation of microsatellite markers for identification of individual Greenshell™ mussels (Perna canaliculus) in a selective breeding programme

A set of 49 microsatellite loci isolated from the endemic New Zealand Greenshell™ mussel, Perna canaliculus, were evaluated for inclusion in a parentage assignment marker suite by assessing their ease of PCR amplification, allele scoring and conformity to Mendelian inheritance in hatchery-produced families. Ten polymorphic loci (mean H_e = 0.78 and polymorphic information content (PIC) = 0.72) were identified as being suitable for parentage assignment. These 10 microsatellite loci gave a combined non-exclusion probability of < 0.001 (probability that an unrelated parent pair will not be excluded from parentage of an arbitrary offspring), based on allele frequencies from 16 broodstock mussels. Simulations predicted an assignment success rate of 99.9% with all 10 loci and > 95% with the best 5 or more loci (mean PIC = 0.84). In actual parentage assignments, 124 offspring from 8 full-sib families were assigned to the correct parent pair with 4 or more loci. We found evidence for null alleles and extensive size homoplasy in many loci, highlighting the importance of thoroughly characterizing and evaluating microsatellite markers prior to parentage assignment and other applications.     

太平洋鳕RAG1和IgM基因荧光定量PCR方法的建立及初步应用

为研究太平洋鳕发育早期特异免疫系统形成的机制,通过RAG1和IgM基因的转录水平衡量特异免疫系统的发育特点.根据GenBank中RAG1和IgM的序列信息,分别设计1对特异引物,从太平洋鳕头肾中扩增得到RAG1和IgM的基因片段.将所获基因片段分别插入到克隆载体pMD18-T中,从而构建太平洋鳕RAG1和IgM基因的质粒标准品.建立并优化太平洋鳕RAG1和IgM基因绝对荧光定量PCR方法.为进一步验证该方法的可靠性,分别利用绝对定量和相对定量检验目的基因在太平洋鳕早期发育过程不同组织内的表达差异.以优化后的绝对荧光定量PCR方法检测不同发育时期太平洋鳕RAG1和IgM的表达情况.结果显示,RAG1的回归方程为y=-3.266x+33.77,回归系数R2=0.996;IgM的回归方程为y=-3.119x +27.61,回归系数R2 =0.998.绝对定量和相对定量结果在基因转录趋势上显现出一致性,即RAG1基因在胸腺和头肾中表达,且在胸腺中的表达量显著高于头肾中的表达量,在肝脏和脾脏中无表达;IgM基因在胸腺、头肾、肝脏和脾脏中均有表达,其中脾脏中表达量最高,其次是头肾.RAG1基因在太平洋鳕发育早期的表达水平很低,到61日龄(days posthatching,dph)至95 dph表达量显著提高;IgM基因在早期表达水平同样很低,到33 dph至61 dph才有明显表达,在95 dph时表达量显著提高.研究表明,本实验方法可靠,特异性较强,可成功对目标基因转录水平进行检测.     

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.     

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.     

Direct and indirect responses to selection on individual body weight in the Pacific oyster (Crassostrea gigas)

Three experiments were performed to examine the heritability of body weight among adult Pacific oysters (Crassostrea gigas) evaluated in Yaquina Bay, Oregon, USA, and to determine if selection on individual body weight could result in changes in offspring survival and yield. The first two experiments utilized midparent-offspring regressions to estimate the heritability (h²) of adult oyster body weight and the coheritability (h_x,y) between adult midparent body weight and offspring performance, including juvenile average body weight, survival and yield as well as adult survival and yield. In Experiment 1 both parents and offspring were evaluated in an “upriver” environment in Yaquina Bay. In Experiment 2 parents were evaluated in a “downriver” environment, while offspring were evaluated in an “upriver” environment. Experiment 3 contrasted average body weight, survival, and yield of offspring (evaluated upriver) derived from three large sires and three small sires mated to the same five females that were chosen at random (all parents evaluated downriver). In Spring 2002, 12 full-sib families from Experiment 1, 19 families from Experiment 2, and 26 families from Experiment 3 were stocked into lantern nets and suspended in Yaquina Bay. Measurements of yield (kg tier^− 1), average body weight (g), and survival (%) were recorded after one and two growing seasons in the field. Heritability estimates for adult body weight at harvest ranged from 0.313 (± 0.083) in Experiment 1 to 0.003 (± 0.073) in Experiment 2. In Experiment 3, average body weight did not differ between offspring derived from large sires and offspring derived from small sires (P = 0.47). Significant negative coheritability estimates were observed between adult midparent body weight and offspring survival in both Experiment 1 and Experiment 2. Significant negative coheritability estimates between adult midparent body weight and offspring yield were observed in Experiment 2 but not in Experiment 1. In Experiment 3, offspring derived from large sires had significantly lower survival and yield than offspring derived from small sires. These results show adult oyster body weight to be heritable but also subject to site-specific adaptation such that selection in the downriver Yaquina Bay environment was ineffective at changing average body weight in the upriver environment. Negative coheritability estimates between performance traits indicate that adult oyster body weight may be a poor indirect measure of oyster yield potential, and that selection solely for increased body weight could lead to a decrease in offspring yield.     

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%.     

The effects of fish hydrolysate (CPSP) level on Octopus maya (Voss and Solis) diet: Digestive enzyme activity, blood metabolites, and energy balance

As has been demonstrated in previous studies, Octopus maya can be fed on artificial diets. In the present study six different diets were assayed. Five diets were designed to test the effect of percentage of inclusion of fish protein concentrate (CPSP: 0, 5, 10, 15, and 20%) and were offered to octopuses as a specifically designed artificial diet. The sixth diet consisted of frozen crab (Callinectes spp) and was used as control diet. Blood metabolites and energy budget of octopuses were evaluated to determine how CPSP levels modulate the digestive capacity and allow retaining energy for growth. Wild animals (316.4 ± 9.8 g) were used in the study. Results showed that CPSP produced a positive specific growth rate (SGR, % day^− 1) with high value in octopuses fed 15% CPSP level. A maximum growth rate of 0.86% day^− 1 was recorded in these animals, a value that is extremely low when compared with the SGR obtained when animals were fed fresh crab (3.7% day^− 1). In general, blood metabolites were affected by diet composition, indicating that some metabolites could reflect the nutritional and/or physiological status of octopus. Preliminary reference values for O. maya fed crab were found for glucose (0.09 ±0.02 mg/ml), lactate (0.004 ± 0.002 mg/ml), cholesterol (0.16 ± 0.02 mg/ml), acylglycerol (0.14 ± 0.01 mg/ml), protein (0.37 ± 0.04 mg/ml), hemocyanin (1.85 ± 0.04 mmol/l), and digestive gland glycogen (1.86 ± 0.3 mg/g). Total energy content can be used as an indicator of tissue metabolic reserves. In the present study, higher energy content in the digestive gland and muscle was observed in octopuses fed crab, followed by animals fed 15% CPSP. Results from the digestive gland indicated that the retained energy derived from glycogen, suggesting that lipids and protein were the main sources of variation linked with energy content. In general, digestive gland proteases activity and trypsin were induced in octopuses fed 15% CPSP. The capacity of O. maya juveniles to adjust their digestive enzymes to different types of food was evidenced. Essential amino acid content (EAA) of the diet was not a limiting factor. When dietary EAA profiles were compared with O. maya EAA profiles, all dietary EAA resulted in a higher concentration than whole body octopus composition. In the present study, all experimental groups ingested between 3300 and 4106 kJ wk^− 1 kg^− 1 without statistical differences among treatments, indicating that experimental diets were as attractive as crab. Differences were recorded in the proportion of absorbed energy (Ab, %) between CPSP-based and crab meat diets, suggesting digestion limitations associated with artificial diets. The present results indicate that the 15% CPSP diet had characteristics that stimulate digestive enzymes and reduce energetic costs associated with its digestion (HiE or SDA), channeling more biomass production than the other experimental diets.     

A microsatellite marker tool for parentage analysis in Senegal sole (Solea senegalensis): Genotyping errors, null alleles and conformance to theoretical assumptions

A set of 50 potential microsatellite loci previously described in several flatfish species was evaluated for designing a suitable tool for parentage assessment in Senegal sole (Solea senegalensis). Low cross-amplification within Pleuronectiformes, even between congeneric species, was achieved. This suggests a low conservation of microsatellite flanking regions in this order regarding other fish orders of similar evolutionary age. Twelve loci showed appropriate amplification and polymorphism to acquire enough statistical power for parentage analysis. All forty-one individuals from one broodstock and 320 offspring were analyzed using this set of markers. The high potential of microsatellites for parentage exclusion (Excl1 = 0.9999; Excl2 = 1) permitted us to trace back all offspring to single pairs of parents. The families obtained were used to analyze the performances of the loci applied and to identify possible sources of error for parentage analysis in Senegal sole. The 2.5% of single mismatches in the families studied were attributed mostly to genotyping errors and to a lesser extent to null alleles. The average mutation rate estimated over 7680 gametes was in the range reported within fish (3.9 × 10^− 4) and did not compromise parentage analysis. All loci conformed to theoretical assumptions (Mendelian segregation, independent segregation), excluding the F13-II8/4/7 and Sol 9A loci, which showed strong genotypic association in the families analyzed. A set of 4 microsatellites (Sol CA13, F13-II8/4/7, Smax-02 and Sse GATA38) was finally selected for parentage evaluation in S. senegalensis combining both enough exclusion potential (> 0.970) and the lowest economic cost.