Efficacy of bithionol as an oral treatment for amoebic gill disease in Atlantic salmon Salmo salar (L.)

This study examined the efficacy of bithionol as an oral treatment for Atlantic salmon Salmo salar affected by amoebic gill disease (AGD). The current commercial management strategy of AGD is a costly 3 h freshwater bath. It is labour intensive and the number of baths needed appears to be increasing; hence, there is an effort to identify alternative treatments. Efficacy was examined by feeding AGD-affected Atlantic salmon twice daily to satiation with bithionol, an antiprotozoal, at 25 mg kg^− 1 feed. Three seawater (35‰, 17 °C) re-circulation systems were used each consisting of three tanks containing 32 Atlantic salmon smolts with an average (± SEM) mass of 90.4 g (± 5.2). Three feeds were examined in the trial including bithionol, plain commercial control and oil coated commercial control. Feeding commenced 2 weeks prior to exposure to Neoparamoeba spp. at 300 cells L^− 1 and continued for 28 days post-exposure. Efficacy was determined by examining gross gill score and identifying percent lesioned gill filaments twice weekly for 4 weeks post-exposure. Bithionol when fed as a two-week prophylactic treatment at 25 mg kg^− 1 feed delayed the onset of AGD pathology and reduced the percent lesioned gill filaments by 53% and halved the gill score from 2 to 1 when compared with both the plain and oil controls during an experimental challenge. There were no palatability problems observed with mean feed intake of bithionol over the trial duration with fish consuming higher levels of the bithionol diet compared to both the oil and plain controls. This study demonstrated that bithionol at 25 mg kg^− 1 feed, when fed as a two-week prophylactic treatment for Neoparamoeba spp. exposure, delayed and reduced the intensity of AGD pathology and warrants further investigation as an alternative to the current freshwater bath treatment for AGD-affected Atlantic salmon.     

Effect of water temperature on the immune response of white shrimp Litopenaeus vannamei to Vibrio alginolyticus

White shrimp Litopenaeus vannamei held in 25‰ seawater at 27 °C or 28 °C were injected with TSB-grown Vibrio alginolyticus at 1 × 10⁴ colony-forming units (cfu) shrimp^− 1 or 1 × 10⁵ cfu shrimp^− 1, and then cultivated onward at water temperatures varying from 20 to 34 °C. Over 24–144 h, mortality of V. alginolyticus-injected shrimp held at 34 °C or 32 °C was significantly higher than that of shrimp held at lower temperatures. In a separate experiment, shrimp held in 25‰ seawater at 28 °C and then cultured onward at 20 to 32 °C were examined for immune parameters at 24–96 h. THC, phenoloxidase activity, respiratory burst, and SOD activity decreased significantly at 24 h after transfer to 32 °C. Shrimp held in 25‰ seawater at 27 °C and then cultured onward at 20 to 34 °C showed a significant reduction in phagocytic activity and clearance efficiency for V. alginolyticus at 24 h after transfer to 34 °C. It was concluded that transfer of shrimp from 27 or 28 °C to higher temperatures (32 and 34 °C) reduced their immune capability and decreased resistance to V. alginolyticus infection.     

Furunculosis in Atlantic salmon (Salmo salar L.) is not readily controllable by bacteriophage therapy

The potential of bacteriophage therapy to control bacterial disease in farmed fish was tested using, as an example, furunculosis of Atlantic salmon, caused by Aeromonas salmonicida subsp. salmonicida.

In vivo testing with Atlantic salmon and rainbow trout (Oncorhynchus mykiss Walbaum) showed no adverse effects, with bacteriophage generally cleared within 96 h of administration by either intraperitoneal (i.p.) injection or oral in-feed.

Juvenile Atlantic salmon were administered a combination of bacteriophage O, R and B (1.9 × 10⁸ pfu fish^− 1) by i.p. injection, after they had been challenged with A. salmonicida subsp. salmonicida 78027, also by i.p. injection. The fish that were injected with bacteriophage immediately after challenge died at a significantly slower rate then those that were either not treated with bacteriophage, or treated 24 h post-challenge. However, the end result (100% mortality) was not affected.

In further experiments the effects of oral (1.88 × 10⁵ pfu g^− 1 fish^− 1 daily for 30 days), bath (1.04 × 10⁵ ml^− 1 daily for 30 days) and i.p. (6.25 × 10⁷ pfu fish^− 1) phage treatment to control furunculosis in experimentally infected Atlantic salmon were compared with antibiotherapy (treatment with 10 mg kg^− 1 bw^− 1 day^− 1 oxolinic acid for 10 days), using an indirect cohabitation challenge. No protection was offered by any of the bacteriophage treatments, compared to the positive challenge group, although significant protection was offered by the oxolinic acid treatment. Analysis of samples taken from the trials demonstrated that bacteriophage were correctly administered to the fish and, on occasion, were isolated from fish that had succumbed to furunculosis. It was also shown that bacteriophage resistant A. salmonicida subsp. salmonicida isolates could be recovered from mortalities in all the treatment groups.

The results suggest that, although there were no safety problems associated with the approach, furunculosis in Atlantic salmon is not readily controllable by application of bacteriophage.     

Metabolic scope, swimming performance and the effects of hypoxia in the mulloway, Argyrosomus japonicus (Pisces: Sciaenidae)

The culture of the mulloway (Argyrosomus japonicus), like many other Sciaenidae fishes, is rapidly growing. However there is no information on their metabolic physiology. In this study, the effects of various hypoxia levels on the swimming performance and metabolic scope of juvenile mulloway (0.34 ± 0.01 kg, mean ± SE, n = 30) was investigated (water temperature = 22 °C). In normoxic conditions (dissolved oxygen = 6.85 mg l^− 1), mulloway oxygen consumption rate (M·o₂) increased exponentially with swimming speed to a maximum velocity (U_crit) of 1.7 ± < 0.1 body lengths s^− 1 (BL s^− 1) (n = 6). Mulloway standard metabolic rate (SMR) was typical for non-tuna fishes (73 ± 8 mg kg^− 1 h^− 1) and they had a moderate scope for aerobic metabolism (5 times the SMR). Mulloway minimum gross cost of transport (GCOT_min, 0.14 ± 0.01 mg kg^− 1 m^− 1) and optimum swimming velocity (U_opt, 1.3 ± 0.2 BL s^− 1) were comparable to many other body and caudal fin swimming fish species. Energy expenditure was minimum when swimming between 0.3 and 0.5 BL s^− 1. The critical dissolved oxygen level was 1.80 mg l^− 1 for mulloway swimming at 0.9 BL s^− 1. This reveals that mulloway are well adapted to hypoxia, which is probably adaptive from their natural early life history within estuaries. In all levels of hypoxia (75% saturation = 5.23, 50% = 3.64, and 25% = 1 .86 mg l^− 1), M·o₂ increased linearly with swimming speed and active metabolic rate (AMR) was reduced (218 ± 17, 202 ± 14 and 175 ± 10 mg kg^− 1 h^− 1 for 75%, 50% and 25% saturation respectively). However, U_crit was only reduced at 50% and 25% saturation (1.4 ± < 0.1 and 1.4 ± < 0.1 BL s^− 1 respectively). This demonstrates that although the metabolic capacity of mulloway is reduced in mild hypoxia (75% saturation) they are able to compensate to maintain swimming performance. GCOT_min (0.09 ± 0.01 mg kg^− 1 m^− 1) and U_opt (0.8 ± 0.1 BL s^− 1) were significantly reduced at 25% dissolved oxygen saturation. As mulloway metabolic scope was significantly reduced at all hypoxia levels, it suggests that even mild hypoxia may reduce growth productivity.     

Dopamine modulates the physiological response of the tiger shrimp Penaeus monodon

Levels of glucose, lactate, pO₂, pCO₂, HCO₃⁻, TCO₂, Na⁺, K⁺, Cl⁻, protein, and oxyhemocyanin in the hemolymph and its osmolality and pH were measured when tiger shrimp, Penaeus monodon (13.5 ± 1.5 g body weight), were individually injected with saline or dopamine at 10^− 8, 10^− 7, or 10^− 6 mol shrimp^− 1. Results showed that hemolymph glucose, lactate, pCO₂, HCO₃⁻, and TCO₂ values increased from 2 to 4 h; hemolymph osmolality, Na⁺, and total protein had increased at 2 h; and hemolymph K⁺ decreased from 2 to 8 h after the dopamine injection. All physiological parameters returned to the control values 4–16 h after receiving dopamine. The dopamine injection also significantly decreased the oxyhemocyanin/protein ratio of P. monodon which occurred at 2 h, resulting from an elevation of hemolymph protein and a slight decrease of oxyhemocyanin. These results suggest that stress-inducing dopamine caused a transient period of modulation of energy metabolism, osmoregulation, respiration, and the acid–base balance in P. monodon in adapting to this environmental stress.     

Physiological changes in rainbow trout held under crowded conditions and fed diets with different levels of vitamins E and C and highly unsaturated fatty acids (HUFA)

Rainbow trout (Oncorhynchus mykiss) maintained in crowded (100 kg m^− 3) and uncrowded (20 kg m^− 3) conditions were fed 42 days with five experimental diets having different levels of vitamin E (25.6 and 275.6 mg kg diet^− 1), C (0 and 1000 mg kg diet^− 1) and HUFA (highly unsaturated fatty acids, 12.5 and 320.5 g kg diet^− 1): −E−HUFA, −E+HUFA, +E−HUFA, +E+HUFA, −C+E+HUFA. Cortisol, plasma metabolites, tissue glycogen, fish composition, and tissue fatty-acid profile were evaluated at the end of the experimental period. In general, no changes in cortisol levels were associated with crowding, although +E+HUFA and −C+E+HUFA fish showed higher levels (mean ± SE, 55.5 ± 11.1 and 78.0 ± 11.3 ng ml^− 1) as a consequence of a possible interaction between chronic crowding and diet composition. Protein and glucose con-centration in plasma displayed no effect of crowding, but liver glycogen showed a general tendency to decrease in −E−HUFA, −E+HUFA, +E−HUFA, +E+HUFA, −C+E+HUFA crowded groups (70.2 ± 2.1, 52.1 ± 2.5, 73.4 ± 7.4, 91.7 ± 3.3, 74.2 ± 8.4 mg g^− 1 tissue, respectively) compared to uncrowded groups (108.9 ± 14.2, 82.7 ± 8.8, 92.4 ± 10.7, 99.1 ± 10.0, 103.5 ± 15.6 mg g^− 1 tissue, respectively), thus proving significant in −E+HUFA fish. Variations in total lipids, triglycerides, total cholesterol and HDL as well as LDL cholesterol in plasma were manifested under crowding conditions, displaying a certain influence of vitamin E and HUFA dietary content. Final body composition, in general, showed no change attributable to fish density, but some differences associated with diet composition were found in lipid and moisture percentages of crowded fish. Liver and muscle fatty-acid profile revealed a clear effect of the dietary lipid source that was more evident in muscle than in liver at normal fish density, and in some cases this effect was modulated by dietary vitamin E and C content and fish-culture conditions.     

An improved enzyme preparation for rapid mass production of protoplasts as seed stock for aquaculture of macrophytic marine green algae

Preparation of protoplasts and their subsequent applications for both basic and applied research of marine macroalgae remains largely under developed due to lack of development of reliable methods with consistent yields of viable protoplasts. An improved enzyme preparation with a single commercial enzyme, e.g. 2% Cellulase Onozuka R-10 in 1% NaCl solution, was developed to produce protoplasts rapidly from different green algal genera of Ulva, Enteromorpha and Monostroma. The simple dissolution of enzyme powder in 1% NaCl resulted in exclusion of 2% Macerozyme R-10 from the mixture consisting of 2% Cellulase Onozuka R-10 with 3% NaCl earlier reported as superior for the same algae. Optimal conditions for the isolation of maximum yields of viable protoplasts were found to be with 2% Cellulase Onozuka R-10 incubated at 20 °C for 2 h in 1% NaCl solution with 0.8 M mannitol adjusted to pH 6.0. The protoplast yield with optimized enzyme mixture was as high as 102.8 × 10⁶ cells g^− 1 f. wt for M. oxyspermum while it was in the range of 74.4–88.6 × 10⁶ cells g^− 1 f. wt thallus for seven species of Ulva, and 82.5–95.4 × 10⁶ cells g^− 1 f. wt for three species of Enteromorpha. The regeneration rate of protoplasts isolated using this method ranged from 89 to 92% with normal morphogenesis. The seeding of nylon threads with isolated protoplasts of M. oxyspermum was successful and after 3–4 weeks the entire frame with nylon threads became thick green in color with tiny germlings in laboratory culture. Thus, the method described in the present study allow for rapid mass production of viable protoplasts that could be potentially used as a source for seed material for mariculture and for other applied phycological research.     

Breeding and juvenile culture of the lined seahorse, Hippocampus erectus Perry, 1810

All seahorse species worldwide have been placed under CITES Appendix II since 2004, because they have been over-exploited for traditional Chinese medicine and aquarium trades. Aquaculture has been recognized as a long-term solution for sustaining the seahorse trade while minimizing wild collection. In this study, we evaluated the breeding and juvenile culture of an important aquarium seahorse species, the lined seahorse Hippocampus erectus, Perry 1810. Pairing, mating and copulation behavior were observed. Gestation time and brood size were 17.33 ± 2.94 days and 272.33 ± 66.45 individuals/brood, respectively. Growth rates differed among juveniles from different broods. Effects of temperature on the growth rates and survivorship of the juveniles during the first two weeks were compared. The highest growth rate and survivorship of the juveniles occurred at 28–29 °C among the temperatures tested (24–33 °C). Growth rate and survivorship of the juveniles during the first 9 weeks at 28 °C were investigated. The final standard length and survivorship of the juveniles were 6.32 ± 0.52 cm and 71.11 ± 10.18%, respectively, and the relationship between the wet weight and the standard length of the juvenile seahorses can be expressed as: W = 0.0034 L^2.5535 (r² = 0.9903, n = 12, P < 0.01). These findings suggest that H. erectus is a good candidate for commercial aquaculture.     

Energy and nitrogen partitioning in 250 g Atlantic cod (Gadus morhua L.) given graded levels of feed with different protein and lipid content

The effects of feed intake level on energy and nitrogen partitioning were studied in juvenile Atlantic cod (250 g) fed two fish meal based diets differing in protein and lipid content (54:31 and 65:16) at 10 °C. Replicate groups of cod were feed deprived for 32 days or fed one of the two diets at 25, 50, 75 or 100% of group satiation for 60 days. Feed intake and oxygen consumption were measured daily and weights and chemical composition of carcass, liver, viscera and whole body were measured at start and end. Diet digestibilities were assessed in a separate experiment.

The whole body and carcass growth rates at a given feed intake did not differ between dietary groups, but the liver grew faster in the fish fed the low protein diet, resulting in higher hepatosomatic indices at the end of the experiment in the groups fed this diet.

The efficiency of utilisation of digestible nitrogen for growth (k_DNg) was higher for the low protein diet (0.73 ± 0.02) than for the high protein diet (0.53 ± 0.05), resulting in higher nitrogen retention at a given nitrogen intake. No difference in percentage nitrogen retention was seen in full-fed fish however (31.2 ± 2.5 and 28.4 ± 1.6% for the low protein and high protein diets, respectively). This can be explained by higher nitrogen intake in the fish fed the high protein diet, resulting in a smaller proportion of the intake being used for maintenance.

There was no difference in energy utilisation between dietary groups. The digestible energy requirement for maintenance (DE_maint) was 53.8 ± 0.9 kJ kg^− 1 d^− 1 (42.3 ± 0.7 kJ kg^− 0.8 d^− 1) and the utilisation efficiency for growth (k_DEg) was 0.80 ± 0.02. The energy retention in full-fed fish was 31.3 ± 3.5 and 31.7 ± 1.0% for the low protein and high protein diets, respectively. The deposited energy was distributed in approximately equal proportions in the liver and carcass, whereas viscera accounted for a minor proportion. At a given energy intake, the fish fed the high protein diet deposited more energy in the carcass and less in the liver than did those fed the low protein diet.     

Characterization of Ca-ATPase activity in gill microsomes of freshwater mussel, Lamellidens marginalis (Lamarck) and heavy metal modulations

Freshwater mussel Lamellidens marginalis showed a Ca²⁺-stimulated ATPase activity in the microsomal fraction of gill. Mg²⁺ was also found to be an activating cation for this ATPase enzyme. Ca²⁺ ATPase showed maximal activity at 40 °C and between pH 7.5–8.0. Substrate specificity of Ca²⁺ ATPase was highest with ATP, followed by GTP and other trinucleotides such as UTP, CTP and ADP also showed some amount of hydrolysis. Ca²⁺ ATPase showed slight inhibition with ruthenium red and sodium vanadate and is insensitive to sodium azide, ouabain, oligomycin B and phenylalanine. Heavy metals like Hg²⁺, Cu²⁺ and Zn²⁺ showed 50% inhibition of Ca²⁺ ATPase activity at concentrations of 0.25 mM, 0.5 mM and 1 mM, respectively whereas Cd²⁺ and Ni²⁺ showed 39% and 28% inhibition of enzyme activity at 1 mM concentrations.     

Purification and partial characterization of Cu, Zn superoxide dismutase from haemolymph of Oriental river prawn Macrobrachium nipponense

The copper plus zinc superoxide dismutase (Cu, Zn-SOD) was purified from haemolymph of the Oriental river prawn, Macrobrachium nipponense and partially characterized. Partial protein precipitation in crude extract was affected by using heat treatment and (NH₄)₂SO₄ fractionated precipitation methods. Fractionation of superoxide dismutase was performed by DEAE-cellulose 32 ion-exchange chromatography and followed by CM-cellulose cation-exchange chromatography. The molecular weight of it was about 66.1 kDa, as judged by SDS polyacrylamide gel electrophoresis. The enzyme was sensitive to cyanide and H₂O₂, and contained 1.08 ± 0.14 atom of copper and 0.98 ± 0.11 zinc per subunit shown in atomic absorption spectroscopy, which revealed that purified SOD was Cu, Zn superoxide dismutase. The purified enzyme had an absorption peak of 269 nm in ultraviolet region and the enzyme remained stable at 25–45 °C within 60 min. But it was rapidly inactivated at higher temperature (50 °C). The activity of purified shrimp Cu, Zn-SOD was remained stable over the range pH 5.8–8.3. Treated with 10 mM mercaptoethanol, the enzyme activity significantly increased. However, the enzyme activity was obviously inhibited by 10 mM CaCl₂, ZnCl₂, SDS, EDTA–Na₂ and 1 mM and 10 mM K₂Cr₂O₇. The results showed that it might be a kind of EC-SOD. And it was the first report of some characterizations of this EC-SOD in M. nipponense.     

Bacterial diversity of tilapia (Oreochromis niloticus) cultured in brackish water in Saudi Arabia

The bacterial flora occurring in brackish pond water, sediment, gills and intestine of healthy tilapia cultured in Saudi Arabia were estimated both quantitatively and qualitatively, and the isolates were identified to genus or species level. Total viable count of bacteria ranged from 1.4±1.5×10³ to 8.6±2.7×10³ cfu ml⁻¹; 1.2±3.1×10⁶ to 7.3±1.1×10⁷ cfu g⁻¹; 8.7±1.9×10⁵ to 2.1±0.9×10⁶ cfu g⁻¹; and 2.8±2.4×10⁷ to 1.0±1.6×10⁸ cfu g⁻¹ in the pond water, sediment, gills and intestine of brackish water tilapia, respectively. In total, 19 bacterial species were identified. The bacteria were predominantly Gram-negative rods (87%). Pond water and sediment bacteria influenced the bacterial composition of gills and intestine of tilapia. In contrast to gill bacteria, more diversification was observed in intestinal bacteria. The predominant (prevalence >10%) bacterial species were Vibrio parahaemolyticus, Vibrio carchariae, Vibrio alginolyticus, Chryseomonas sp., Vibrio vulnificus, and Streptococcus sp. in all the populations with the exception of the sediment population where Streptococcus sp. was replaced by Shewanella putrefaciens. Vibrio spp. (58% of the total isolates) dominated the total bacterial population.     

Ammonia and pH effects on some metabolic parameters and gill histology of silver catfish, Rhamdia quelen (Heptapteridae)

The aim of the present study was to assess the effect of ammonia exposure at different pH on survivorship and metabolic parameters in the liver, muscle and gill histology of silver catfish (Rhamdia quelen). The 96 h-LC50 of un-ionized ammonia (mg L^− 1) at pH 6.0, 7.5 and 8.2 were: 0.44 (C.I. 0.38–0.49), 1.45 (C.I. 1.25–1.65) and 2.09 (C.I. 1.85–2.36), respectively. Survival of juveniles exposed to different ammonia levels was altered by pH, and fish exposed to all ammonia levels and different pH showed muscle glucose, muscle and liver glycogen reduction. Liver glucose and muscle and liver lactate levels increased in all fish exposed to ammonia as compared to the control. Exposure to waterborne ammonia increased total ammonia levels in both tissues and also induced gill epithelium damages such as lamellar fusion and edema as compared with controls at different pH. Silver catfish exposed to pH 6.0 and different NH₃ levels presented significantly higher hepatic glucose and protein levels when compared to those maintained at low NH₃ levels. Juveniles exposed to NH₃ levels at pH 7.5 and 8.2 showed lower hepatic protein levels compared to those maintained at low NH₃ levels. These parameters are indicative of pH dependence on ammonia toxicity in silver catfish. The metabolic parameters and gill histology may be used as early indicators of ammonia toxicity in silver catfish.     

Influence of different food sources on the initial development of sexual recruits of reefbuilding corals in aquaculture

In coral aquaculture, sexual reproduction increasingly plays an important role for serving trade and reef restoration purposes. However, until coral juveniles reach a semi-stable size which makes them less vulnerable against algal growth and sedimentation, high mortality rates may occur in the first several weeks to months after settlement. In the present study, the influence of several food sources on the growth and survival of newly settled primary polyps was studied under laboratory conditions for 5 months. In order to estimate effects on specimens of both reproductive modes, experiments were carried out with the brooder, Favia fragum, and with the broadcast spawner, Acropora tenuis. Primary polyps kept in 2-liter aquaria were daily fed with freshly hatched Artemia salina, the micro algae Phaeodactylum tricornutum and a commercially available dry food (Nori Micro, Zoolife®), respectively, at various concentrations. Growth rates in both species were significantly higher in the Artemia treatment with maximum rates of 9.4 ± 4.9 mm² (mean ± S.D.; d = 5 months) for F. fragum and 26.8 ± 10.3 mm² (d = 5 months) for A. tenuis compared to all other treatments and the control (no additional food). Survival in F. fragum was overall higher than 60% in all treatments with maximum rates of 85.0 ± 12.6% at the highest Artemia concentration. Survival rates in A. tenuis ranged from 28.9 ± 4.7% (lowest Nori concentration) to 86.2 ± 5.9% (medium Artemia concentration). The present study shows that Artemia nauplii may greatly enhance the growth and partly enhance the survival of early sexual recruits which may significantly help to more rapidly overcome the early and most fragile post-settlement stages. As a consequence, the residence time for sexual coral recruits in cost- and labour-intensive hatcheries may be greatly reduced.     

Streptococcus iniae infection and tissue distribution in hybrid striped bass (Morone chrysops×Morone saxatilis) following inoculation of the gills

This study was designed to test the possibility that Streptococcus iniae enters through the gills and causes infection in hybrid striped bass. To determine the dose response, four groups of fish were inoculated with S. iniae via the gills with a dose of 5.0×10⁵, 2.6×10⁶, 5.0×10⁶, or 1.0×10⁸ CFU/fish. One group of fish was inoculated with tryptic soy broth (TSB) via the gills to serve as controls. The cumulative percent mortality was 13%, 27%, 100% and 100% for 5.0×10⁵, 2.6×10⁶, 5.0×10⁶ and 1.0×10⁸ CFU/fish, respectively. We also examined the tissue dissemination of S. iniae at 0.5, 4, 8, 12, 24, 48 and 72 h after experimental gill inoculation. Fish were inoculated with 2.6×10⁶ or 5.0×10⁶ CFU/fish, which caused low and high mortality, respectively. Within 48 h, fish inoculated with the 2.6×10⁶ dose were culture positive on the gill surface, blood of the first and second gill arches, blood of the third and fourth gill arches and the nares. However, for the dose of 5.0×10⁶ CFU/fish, S. iniae was also isolated from the olfactory, optic and cerebellum regions of the brain, eye, head kidney, trunk kidney, spleen and liver at 48 h. For the 2.6×10⁶ dose, S. iniae was not isolated until 48 h post-inoculation, but was isolated at 12 h for the 5.0×10⁶ dose. The results of this study indicate that S. iniae can enter hybrid striped bass through the gills. However, mortality at similar S. iniae doses was lower than we previously observed by inoculation of the nares.     

Evidences of the intertidal red alga Grateloupia turuturu in turning Vibrio parahaemolyticus into non-culturable state in the presence of light

Grateloupia turuturu, previously known as Grateloupia doryphora, has been widely reported to be an invasive algal species. There are no studies to relate the impact of its existence on its surrounding environment. In this paper, we present our results to show that about 70% of individuals collected from the field could turn Vibrio parahaemolyticus into non-culturable state on both selective (TCBS) and non-selective (2216E) culture medium in 24 h in the presence of light in live algal culture. Total bacteria counts on TCBS and 2216E plates dropped from the initial 565 (± 174) and 1192 (± 60) cfu ml^− 1 respectively to zero in 24 h. This effect disappeared when the alga was grown in darkness. The same effect was not found in two other intertidal macroalgae Laminaria japonica and Palmaria palmata. Further tests showed that the settlement ability of bacteria in seawater was impaired significantly in the presence of this alga in comparison with three other algal species.     

Serotonin induces ovarian maturation in giant freshwater prawn broodstock, Macrobrachium rosenbergii de Man

This study investigated the effects of serotonin (5-hydroxytryptamine or 5HT) on ovarian development in Macrobrachium rosenbergii de Man. Adult female prawns at the ovarian stage I (spent) were injected with 5HT at 1, 5, 10, 20 and 50 μg g^− 1 body weight (BW) intramuscularly on days 0, 5 and 10, and sacrificed on day 15. The doses as related to the effect could be categorized into three levels: low (1 and 5 μg g^− 1 BW of 5HT), medium (10 and 20 μg g^− 1 BW of 5HT) and high (50 μg g^− 1 BW of 5HT). The low-dose, especially at 1 μg g^− 1 BW, caused prawns to exhibit a significant increase in ovarian index (ovarian weight/body weight × 100) (5.79 ± 0.09%) as compared to the control (1.49%). The ovaries of most of these prawns could develop to stage IV (mature) and contained synchronously mature oocytes while most of the control ovaries remained at stage I and II (proliferative), and contained only oogonia to previtellogenic (Oc1, Oc2) and early vitellogenic oocytes (Oc3). The medium- and high-dose treated prawns exhibited ovaries that could reach stages III and IV and contained various types of oocytes of different maturity. Pretreatment with 5HT receptor antagonist, cyproheptadine (CYP), at 10 μg g^− 1 BW before 5HT injection significantly suppressed the effect of 5HT. Intramuscular injection of the 5HT-primed thoracic ganglion culture medium into CYP-pretreated prawns resulted in the increase of ovarian index about 5–6 times more than in the control, and in the groups injected with 5HT-primed media from muscle strip, eyestalk and brain. The ovaries of most prawn could develop up to stage IV and contained synchronously developed vitellogenic (Oc4) and mature oocytes (Oc5). These findings suggest that 5HT indirectly induces ovarian development and oocytes maturation in M. rosenbergii, probably via a putative ovarian stimulating factor released from the thoracic ganglia.     

Spawning, early development and first feeding in the gobiid fish Priolepis nocturna

The reproductive behavior, embryonic development and early larvae of Priolepis nocturna are described. Three pairs of P. nocturna began spawning 41 days after acquisition and maintained a 5–10 day spawning cycle lasting beyond several months. Spawning was initiated by the female who signaled her readiness to spawn by displaying to the male. Egg clutch size averaged 1578 ± 51.23 eggs and ranged from 268 to 3121. Egg length averaged 0.82 ± 0.01 mm total length (TL) and ranged from 0.75 to 0.90 mm. Egg width averaged 0.51 ± 0.51 mm total width (TW) and ranged from 0.49 to 0.52 mm. Fertilized eggs were ovoid in shape and attached to the ceiling of provided shelters via adhesive filaments at the proximal end. Hatching rates averaged 97.3 ± 0.51% and ranged from 91.9 to 99.8%. Larvae measuring 1.89 ± 0.04 mm TL hatched 121 ± 0.5 h post fertilization and did not rotate position prior to hatching. Skeletal elements of the chondrocranium were simplistic and dominated by the hyoid, hyomandibulosymplectic cartilage, ethmoid and Meckel's cartilage in first feeding larvae. No elements were added to the cranial architecture by 5 days post hatch (DPH) when larvae measured 2.05 ± 0.04 mm TL. First feeding larvae consumed only dinoflagellates and tintinnids suggesting that feeding was constrained by a poorly developed feeding mechanism. Embryology and larval development are described to 5 DPH.     

Extenders and cryoprotectants for cooling and freezing of piracanjuba (Brycon orbignyanus) semen, an endangered Brazilian teleost fish

The aim of this study was to develop a protocol for semen storage of piracanjuba (Brycon orbignyanus) by both cool storage at 4 °C and cryopreservation at − 196 °C. Semen was diluted in some fish semen extenders (Exp. 1) or in extenders combined with the antibiotic gentamycin sulfate (Exp. 2) and stored at 4 °C. Sperm motility was estimated every 24 h. Then, the effects of egg yolk (0 and 5%), cryoprotectants (dimethyl sulphoxide — DMSO, methanol, and methylglycol) and extenders (NaCl 154 mM, BTS™ Minitub and M III™ Minitub) on semen cryopreservation were evaluated (Exp. 3). Semen was added to each of eighteen cryosolutions (2 yolk concentrations × 3 cryoprotectants × 3 extenders), aspirated into 0.5-mL straws, frozen in nitrogen vapor (Taylor-Wharton, CP 300, “dry shipper”) and stored at − 196 °C. Sperm motility was evaluated after thawing at 60 °C-water bath for 8 s. The three cryosolutions that produced the highest post-thaw sperm motility were used again to freeze semen. Post-thaw semen quality was then evaluated under three tests: sperm motility, the percentage of live spermatozoa and hatching rate (Exp. 4). Piracanjuba semen diluted (1:10 total volume) in NaCl 200 mM or in Saad solution (NaCl 200 mM, Tris 30 mM) maintained motility above 35% for as long as 7 days, at 4 °C. Motility of only 7% was observed on undiluted semen after 3 days at 4 °C. There was neither beneficial nor detrimental effect of gentamycin on sperm motility at 250 μg/mL. Egg yolk addition to the cryosolution was beneficial in samples cryopreserved in NaCl 154 mM and in M III™, but detrimental for samples cryopreserved in BTS™. Methylglycol was the most effective cryoprotector compared to DMSO and methanol. Motility and percentage of live spermatozoa were similar among semen cryopreserved in NaCl–yolk, M III™–yolk and BTS™, all containing 10% methylglycol, but lower than fresh control. Hatching rates of eggs fertilized with sperm cryopreserved in NaCl–yolk or BTS™ were higher than for eggs fertilized with sperm cryopreserved in M III™–yolk, but lower than control fertilizations. The semen cryopreservation protocols developed here will be used to set up a gene bank for endangered piracanjuba populations.     

Growth efficiency of juvenile barramundi, Lates calcarifer, at high temperatures

Temperature is recognized to be the most important environmental factor affecting growth in fish. Barramundi are cultured over a wide range of temperatures some of which approach the upper thermal tolerance for this species. A growth trial was conducted on juvenile barramundi to examine the effects of high temperatures ranging from the minimum optimal temperature (27 °C) for growth efficiency to the extreme upper thermal limits (39 °C) for feed intake, growth and growth efficiency. Juveniles (4.87 ± 0.32 g) were held at four different temperatures 27, 33, 36 and 39 °C and fed twice daily to satiation (503.5 g kg^− 1 crude protein, 182.5 g kg^− 1 lipid, 150.1 g kg^− 1 ash, 20.52 GE MJ kg^− 1). Feed intake (g·day^− 1) and SGR (%·day^− 1) increased with increasing temperature up to 36 °C. At 39 °C feed intake, growth, feed efficiency ratio, protein efficiency ratio and productive energy value were significantly lower than at the other temperatures. This demonstrates that growth was optimized at temperatures from 27 to 36 °C and that barramundi have a much wider range for maximum growth efficiency than previously thought.