Evaluation of the formulated diet MySpat for feeding hatchery‐reared spat of the green‐lipped mussel,Perna canaliculus (Gmelin, 1791)

As with most bivalves, a high cost component of hatchery and nursery culture of the green‐lipped mussel (Perna canaliculus) is the raising of live micro‐algae feeds. To remedy this, artificial feeds have been developed, however, these have had mixed success for this species. In this experiment, we tested the formulated diet MySpat (sourced from INVE Aquaculture) on P. canaliculus spat of approximately 1 mm in size. Performance measures under study were mussel spat growth and survivorship after 21 days feeding on varying proportions of MySpat. The experimental diets tested, consisted of a mix of T‐Isochrysis galbana, Pavlova lutheri and Tetraselmis suecica micro‐algae that were replaced with increasing proportions of MySpat, i.e. 25%, 50%, 67%, 75%, 90% and 100% MySpat replacement. Our results revealed no significant increase in spat mortality with increasing MySpat diet levels, and growth was equivalent to the micro‐algae only dietary control in all diets except the 90% and 100% MySpat replacement diets. Such results highlight the role MySpat could have for P. canaliculus hatchery operations, particularly with regard to reducing the costs of raising micro‐algae when producing seed for industry. A greater understanding of the particle selection process of this species would be beneficial for future studies seeking to employ artificial diets.     

The use of diatom Skeletonema costatum on aquaculture‐produced purple sea urchin (Paracentrotus lividus) larvae and post‐larvae diet

The quality of the microalgae provided on Paracentrotus lividus larvae rearing is a primordial factor having a direct (nutritional properties) and indirect (water quality) impact on growth, competence and survival. Skeletonema costatum is a diatom commonly used in the bivalve cultivation. However, the use of this diatom in P. lividus larval cultivations is poorly known. The Rhodomonas spp. is a microalgae commonly used in sea urchin larvae culture. Three different diets were tested on P. lividus larvae and post‐larvae cultivation (D1—Rhodomonas marina, D2—S. costatum, D3—mixture of both algae). Larvae fed with the D2 diet (55.8%) and D3 (39.9%) had a survival at 15 DAH higher than D1 (5.5%). The low survival in D1 could be due to the higher microbiological load on microalgae (Vibrio alginolyticus and V. pectenicide). Larvae fed with S. costatum (D2) showed a lower development than other diets. The competency index was lower for larvae fed with the D2. These results show that microalgae diversified diets contribute to a better development of P. lividus larvae. During the settlement and post‐settlement phase, there was also a lower growth of the sea urchin fed with the D2 and a higher survival for D3.     

Two pathogens of Greenshell™ mussel larvae, Perna canaliculus: Vibrio splendidus and a V. coralliilyticus/neptunius-like isolate

Bacterial pathogens of Greenshell™ mussel (GSM) larvae can cause batch losses during hatchery production. Twenty-two isolates were screened using a larval bioassay. Two strains were identified as potential pathogens. Phenotypic identification of these strains revealed two non-reactive Gram-negative, oxidase positive rods. Sequencing of the 16S rRNA gene identified Vibrio splendidus and a V. coralliilyticus/neptunius -like isolate as pathogens of GSM larvae, with an ability to cause 83% and 75% larval mortality in vitro , respectively, at a concentration of 10² CFU mL⁻¹. Histopathology indicated that the route of infection was via the digestive system. Using healthy larvae as target hosts, Koch's postulates were confirmed for the two isolates. This is the first report on pathogens of GSM larvae.     

Assessment of temperature or salinity effects on larval development by catecholamine‐induced metamorphosis of hatchery‐reared flat oyster,Ostrea angasi (Sowerby 1871) larvae

A need to improve larval rearing techniques led to the development of protocols for catecholamine‐induced settlement of flat oyster, Ostrea angasi, larvae. To further refine these techniques and optimize settlement percentages, the influence of salinity or temperature on development of O. angasi larvae was assessed using epinephrine‐induced metamorphosis. Larvae were reared between salinities of 15–35 and temperatures between 14.5 and 31°C. The greatest percentage survival, growth, development occurred when larvae were reared between 26 and 29°C and between salinities of 30 and 35. Larvae reared outside this salinity and temperature range exhibited reduced growth, survival and/or delayed development. Short‐term (1 h) reduction in larval rearing temperature from 26°C to 23.5°C significantly increased larval metamorphosis without affecting larval survival. Short‐term (1 h) increase in larval rearing temperature from 26°C to 29 and 31°C decreased larval survival and metamorphosis. To ensure repeatability in outcomes, tests showed that larvae sourced from different estuaries did not vary significantly in their metamorphic response to short‐term temperature manipulation and epinephrine‐induced metamorphosis.     

Effects of a mix of Bacillus sp. as a potential probiotic for Florida pompano,common snook and red drum larvae performances and digestive enzyme activities

This study examined the effect of a commercial mix of Bacillus sp. on survival, growth and digestive enzyme activities of Florida pompano, red drum and common snook. Larvae were fed either live feed enriched with Algamac 3050 (Control), Algamac 3050 and probiotics (PB), or the previous diet combined with a daily addition of probiotics to the tank water (PB+). Survival was not affected by the treatments for any of the species. At the end of the pompano and snook trial, standard lengths of larvae from the PB and PB+ treatments were significantly greater than for the control larvae. Microbiological analyses were performed at the end of the pompano trial, and numbers of presumptive Vibrio were not a concern in the system. For both pompano and snook, trypsin‐specific activity was higher in PB and PB+ larvae compared with the control larvae. Similarly, alkaline phosphatase activity was higher for the pompano larvae fed the PB and PB+ treatments and for the snook larvae fed the PB+ treatment compared with the control larvae. This experiment suggests that a mix of Bacillus sp. can promote growth through an early maturation of the digestive system during the early larval stages of pompano and snook.     

Characterization of digestive enzymes protease and alpha‐amylase activities in the thick‐lipped grey mullet (Chelon labrosus,Risso 1827)

Activities of pepsin, trypsin, chymotrypsin, alpha‐amylase and lipase, as well as their optima and stability to pH and temperature, were determined in digestive extracts of thick‐lipped grey mullet, Chelon labrosus, of three different sizes: Group 1 (45.2 ± 3.0 g), Group 2 (180.9 ± 4.2 g), and Group 3 (328.5 ± 43.3 g). SDS‐PAGE zymograms were also used to assess the role of serine proteases in the digestive tract of C. labrosus. On the other hand, possible changes in the digestive enzyme profile of C. labrosus during development were observed, with a comparatively lower pepsin activity, higher activities of alkaline proteases and alpha‐amylase and no lipase activity recorded in pre‐adult specimens. It is suggested that these variations are linked to the changes in diet composition with age, moving from a partly carnivorous to a more herbivorous feeding habit.     

Pelagic larval duration of three amphidromous Sicydiinae gobies (Teleostei: Gobioidei) including widespread and endemic species

Sicydiinae species have an amphidromous life cycle during which they undergo a pelagic larval phase allowing them to disperse through the ocean and to recruit in distant island rivers. Hypotheses for the differences observed in dispersal abilities between species include the variation in pelagic larval duration (PLD). However, the implication of the PLD as a proxy for explaining the dispersal ability of a species is not clear in the Sicydiinae subfamily. In this study, otolith microstructure of three Sicydiinae species was analysed. One of these species, Sicyopus zosterophorum, has a widespread distribution in the West Pacific area, whereas the other two species, Smilosicyopus chloe and Akihito vanuatu, are endemic to New Caledonia and to Vanuatu, respectively. Deposition of the daily growth increments on the otoliths of S. zosterophorum was validated using an alizarin complexone time marking technique. We estimated the PLD for the three species by counting the number of growth increments from the core to the metamorphosis check mark, and it was shorter than the one of previous studies on Sicydiinae species. The PLD of the widespread species, S. zosterophorum (54.6 ± 5.6 days), was similar to those of the endemic species, S. chloe (53.6 ± 5.7 days) and A. vanuatu (55.4 ± 7.5 days). Here, we show that in contrast to the most diverse Sicydiinae genus, Sicyopterus, the PLD could not explain endemism, and we must take into account other elements to explain the differences observed in the distribution range.     

Evaluation of frozen Umbrella‐stage Artemia as first animal live food for Litopenaeus vannamei (Boone) larvae

An alternative larval shrimp feeding regime, in which umbrella‐stage Artemia were constituting the first zooplankton source was evaluated in the culture of Litopenaeus vannamei. In a preliminary experiment, umbrella‐stage Artemia were fed to larvae from zoea 2 (Z2) to mysis 2 (M2) stages to identify the larval stage at which raptorial feeding starts and to determine daily feeding rates. The following experiment evaluated the performance of two feeding regimen that differed during the late zoea/early mysis stages: a control treatment with frozen Artemia nauplii (FAN), and a treatment with frozen umbrella‐stage Artemia (FUA). The ingestion rate of umbrella‐stage Artemia increased from nine umbrella per larvae day^?1 at Z2 stage to 21 umbrella per larvae day^?1 at M2. A steep increase in ingestion and dry weight from Z3 to M2 suggests a shift to a raptorial feeding mode at the M1 stage. Treatment FUA exhibited a significantly higher larval stage index (P < 0.05) during the period that zoea larvae metamorphosed to the mysis stage, and a higher final biomass, compared with treatment FAN. Based on these results and on practical considerations, a feeding regime starting with umbrella‐stage Artemia from Z2 sub‐stage can be recommended for L. vannamei larvae rearing.     

Biochemical and fatty acid dynamics during larval development in the razor clam Ensis arcuatus (Bivalvia: Pharidae)

This article reports on the changes in gross biochemical and fatty acid composition of the razor clam Ensis arcuatus larvae throughout development. Protein was the largest component of dried larval tissues. The energy required for embryogenesis in E. arcuatus oocytes was obtained from stored proteins and carbohydrates, while total lipids increased significantly. Lipids and carbohydrates have an important role as energy contributors from day 1 to day 8. During larval development the strategy was to indistinctly store energy reserves (protein, lipid and carbohydrate) to surpass metamorphosis. During embryonic development there was a gain in fatty acids of neutral and polar lipids. A depletion of fatty acids in neutral lipids was observed from day 1 to day 8 in E. arcuatus larvae, mainly due to the decrease in 16:0 and EPA. NMID fatty acids were present in amounts similar to those of some PUFAs in polar lipids, especially 22:2NMID.     

Effect of live algae used as green water on survival,growth, behaviour,ontogeny and bacterial profile of lobster larvae (Homarus americanus Milne Edwards)

Green water is a technique commonly used in aquaculture, that consists of adding live algae in the water culture and its benefits have been shown for several species. Several hypotheses exist to explain the benefits of green water: increase in nutritional value; action as a probiotic; increase in contrast to reveal preys for larvae; or increase in predator behaviour of larvae. Green water produced with a mix of strains Isochrysis galbana and Chaetoceros muelleri (50:50, cells:cells) applied in four different ways was tested. The survival and the growth of American lobster (Homarus americanus) between stage I and stage IV post‐larvae were not affected by the addition of live algae. The lipid classes were not affected by the addition of algae and limited variation was observed in the fatty acids and bacterial profiles. Furthermore, the green water techniques had a limited effect on the behaviour of post‐larvae stage IV lobster at releasing. Behaviour was mostly affected by the age of post‐larvae. The bacteria Lewinella sp., Leucothrix sp. and Thiothrix sp. appeared to represent a common and core component of Stage IV lobster post‐larvae microflora. The results show that the algae do not increase either nutritional value or feed intake of the lobster larvae. Probiotic effect may be more important when larvae are raised in a close system where potential bacterial pathogens could have more chances to colonize the culture. Also, the dark green colour of the larval tank used in this study may have mimicked the effect of green water in the control group. Biochemical results suggest that dietary supplementation with phospholipids and DHA is needed in a lobster hatchery using frozen Artemia and our open formula Dry mix.     

Allometric growth patterns and development in larvae and juveniles of thick‐lipped grey mullet Chelon labrosus reared in mesocosm conditions

Allometric growth and ontogeny were studied in thick‐lipped grey mullet Chelon labrosus reared in mesocosms from 1 to 71 day post hatching (dph). Multivariate allometric analysis of morphometric growth distinguished three distinct developmental stanzas separated by two morphometric metamorphosis lengths (L_m1 = 4.46 ± 0.06 mm; L_m2 = 28.56 ± 1.04 mm). Body mass growth also showed three distinct episodes separated by two inflections, correlated with morpho‐functional changes. First episode concerned pre‐flexion larvae and ended around 4.5 mm‐L_T (14‐dph), coinciding with estimated L_m1. It was distinguished by reduced growth, but intense morphogenesis and differentiation processes. Organogenesis and allometric changes indicated that development priorities concerned feeding efficiency, by improving detection ability (sensory system development), ingestion capacity (head growth) and assimilation performance (digestive system differentiation), together with respiration efficiency (gill development). Second episode concerned post‐flexion larvae and, ended around 8.6 mm‐L_T (25‐dph). It was distinguished by fast growth of trunk and tail, acquisition of adult axial muscle distribution and completion of gill filament development, improving locomotion and oxygenation performances. It corresponded to transition towards metamorphosing stage as indicated by later isometric growth, musculature maturation and acquisition of juvenile phenotype. Metamorphosis seemed to end at L_m2, suggesting to avoid zootechnic handling before this size.     

The importance of dietary HUFA for meagre larvae (Argyrosomus regius; Asso, 1801) and its relation with antioxidant vitamins E and C

Despite the interest of meagre (Argyrosomus regius) as a fast‐growing candidate for Mediterranean aquaculture diversification, there is a lack of information on nutrition along larval development. Importance of highly unsaturated fatty acids (HUFA) and the antioxidant vitamins E and vitamin C has not been investigated yet in this species. Six diets with two levels of HUFA (0.4% and 3% dw), two of vitamin E (1500 and 3000 mg kg^?1) and two of vitamin C (1800 and 3600 mg kg^?1) were fed to 15 dah meagre larvae. Larval growth in total length and dry body weight was significantly lowest in larvae fed diet 0.4/150/180 and showed few lipid droplets in enterocytes and hepatocytes and lower HUFA contents than the initial larvae. Increase in dietary HUFA up to 3%, significantly improved larval growth and lipid absorption and deposition. Besides, among fish fed 3% HUFA, increase in vitamin E and vitamin C significantly improved body weight, as well as total lipid, 22:6n‐3 and n‐3 fatty acids contents in the larvae. Thus, the results showed that 0.4% dietary HUFA is not enough to cover the essential fatty acid requirements of larval meagre and a high HUFA requirement in weaning diets is foreseen for this species. Besides, the results also pointed out the importance of dietary vitamin E and C to protect these essential fatty acids from oxidation, increase their contents in the larvae and promote growth, suggesting high vitamin E and C requirements in meagre larvae (higher than 1500 and 1800 mg kg^?1 for vitamin E and vitamin C respectively).     

Efficacy of single and multiple doses of fenbendazole against gill parasites (Dactylogyrus sp.) of Labeo rohita (Hamilton, 1822) and its physio‐metabolic effects on the fish

The study was conducted to investigate the efficacy and physio‐metabolic responses of single and multiple doses of fenbendazole (FBZ) in Labeo rohita against gill parasite (Dacylogyrus sp.). Two Isoenergetic (395 Kcal DE/100 g) and isonitrogenous (35% CP) purified diets viz. control (without FBZ) and treatment (0.2% FBZ) were prepared. The efficacy of FBZ was measured after administration of single dose (20 mg/kg b. wt.) on 0th day and multiple doses of FBZ, that is, 20 mg/kg b. wt. on 0th, 3rd and 7th day through diet. Although oral administration of FBZ at single dose could significantly reduce the parasite in fish in comparison with the control group, 100% efficacy of the drug was not achieved up to 15th day of treatment. On the other hand, multiple doses of FBZ could show almost 100% efficacy on 15th day in terms of elimination of parasites from gill. The drug induced oxidative stress in examined fish. There was no significant change in activities of hepatic and muscle aspartate aminotransferase and alanine aminotransferase of L. rohita up to 15th day after oral administration of FBZ at single dose. But both single and multiple doses of FBZ could significantly enhance the hepatic superoxide dismutase, catalase and glutathione‐S‐transferase activities in fish up to 7th day of drug administration and significantly decreased thereafter up to 15th and/or 30th day. It is concluded that oral administration of FBZ at multiple doses (20 mg/kg b. wt.) on 1st, 3rd and 7th day could be effective against Dactylogyrus sp. infection in L. rohita.     

Microbial communities associated with early stages of intensively reared orange‐spotted grouper (Epinephelus coioides)

The gut microbiota plays key roles in the health and general welfare of fish larvae, the present study characterized the bacterial communities associated with grouper Epinephelus coioides larvae during a period of 22 days post hatch (DPH) in an intensive hatchery using both cultivation‐based and cultivation‐independent approaches. Both approaches confirmed that bacteria were present in the gut of larvae before and after the onset of exogenous feeding, and the number of cultiviable bacteria increased gradually from 2 DPH to 22 DPH. A more complex bacterial profile was present in larvae fed fertilizer oyster eggs for 4 days (8 DPH), probably as a result of the onset of exogenous feeding. Interestingly, similar internal microbiota were observed in larvae fed fertilized oyster eggs for 4 days (8 DPH) and rotifers for 2 weeks (22 DPH), although different microbial communities were present in the two feeds. This might suggest that the gut environment of E. coioides larvae selects for a common microbiota, which is more closely related with the rearing water than the two feeds. Therefore, bacterial community of the rearing water may play a critical role in the establishment of gut microbiota of fish larvae and more attention should be paid to its practical modulation by using probiotics. In addition, some potentially beneficial bacteria, such as Lactococcus spp., were the major components of the microbiota associated with fertilized oyster eggs, while these bacteria were not detected in larvae samples.     

Effects of black soldier fly (Hermetia illucens) larvae meal protein as a fishmeal replacement on the growth and immune index of yellow catfish (Pelteobagrus fulvidraco)

A 65 days feeding trial was conducted to evaluate the potential of black soldier fly larvae (BSFL) as a fish meal (FM) protein alternative in the diet of yellow catfish. Fish meal protein was replaced with BSFL meal protein at 0% (control), 13%, 25%, 37%, 48%, 68%, 85% and 100%. Compared with the control group, increased growth performances (e.g. weight gain rate increased by 21.7%) and immune indexes (e.g. serum lysozyme activity increased by 6.8%) of yellow catfish fed with diets in which a maximum of 48% FM protein was replaced by BSFL meal protein was determined. The diet in which 25% FM protein was replaced by BSFL meal protein resulted in the greatest growth performances (e.g. weight gain rate increased by 29.1%) and immune indexes (e.g. serum lysozyme activity increased by 31.9%) as well as the lowest feed conversion ratio (0.9) among all diets tested. No significant differences in survival rate, body indexes or composition were determined among all treatments. Therefore, BSFL meal protein has the potential to partially replace FM protein in the diet of yellow catfish, and it may also enhance the immunocompetence of the fish.     

Effect of dietary protein levels on growth and nutrient utilization of fringe‐lipped carp,Labeo fimbriatus (Bloch) fingerlings

Effect of various dietary protein levels on growth and nutrient utilization were studied in fringe‐lipped carp, Labeo fimbriatus fingerlings for 60 days. Five practical diets containing graded protein levels of 200, 250, 300, 350 and 400 g kg^?1 with respective digestible protein (DP) contents of 192.4, 244.5, 291.6, 339.4 and 391.4 g kg^?1 were evaluated as five treatments, T1, T2, T3, T4 and T5, respectively, in triplicate. Each experimental tank (80‐L) contained eight fingerlings (4.9 ± 0.1 g) and was subjected to continuous aeration and 25% water replenishment daily. The fish were fed two times daily at 8:00 and 14:00 h to satiation. Significantly higher (P < 0.05) absolute growth and thermal growth coefficient, and lower feed conversion ratios (FCR) were observed in T2–T4 than T1 and T5. Protein efficiency ratio (PER) and protein productive values (PPV) were highest in T2 and lowest in T5. Significantly higher (P < 0.05) apparent protein digestibility was perceptible in T2–T4. While specific amylase activity declined linearly with increasing DP : DE, the protease, trypsin, aspartate aminotransferase and alanine aminotransferase established polynomial relationship. Based on live weight gain, PER and PPV fitted to quadratic model optimum DP levels were estimated as 301.4, 260.0 and 273.0 g kg^?1, respectively.     

Evaluation and characterization of a novel probiotic Lactobacillus pentosus PL11 isolated from Japanese eel (Anguilla japonica) for its use in aquaculture

In the present study, a potential Lactobacilli probiotics were isolated from Japanese eels (Anguilla japonica) and characterized and evaluated for their possible use in eel farming. Sixteen Lactobacilli were isolated from intestines of Japanese eels, using selective media. The lactobacilli strains (represented as PL1 to PL16) were screened by their ability to produce digestive enzyme. Among these, three strains (PL11, PL13 and PL16) producing four digestive enzymes (amylase, cellulase, protease and phytase) simultaneously were characterized further using API ZYM kit. From these, PL11 (Lactobacillu (L.) pentosus) was identified as potential probiotics candidate producing 15 enzymes among 20 tested. Further examination of biological activities of PL11 revealed tolerance against pH, artificial bile juice and antibacterial activity against several fish pathogenic bacteria. The in vitro competitive exclusion assay also revealed 88.4% reduction in adhesion of fish pathogen (Edwardsiella tarda) by PL11 to host intestinal mucus. In vitro incubation of Japanese eel foregut with Baclight‐labelled PL11 showed colonization of the enterocyte surface by confocal and scanning electron microscopy. In summary, PL11 isolated from eels could serve as a potential probiotics with acid and bile tolerance, production of digestive enzymes, antibacterial activity and inhibition of fish pathogen adhesion to intestinal mucus.     

Ovoviviparously produced Artemia nauplii are a suitable live food source for the larvae of the African catfish (Clarias gariepinus: Burchell, 1822)

Brine shrimp Artemia, the most common live food organism used in larviculture, can reproduce either oviparously (production of dormant cysts) or ovoviviparously (direct production of nauplii), depending on environmental conditions. Ovoviviparous Artemia nauplii have seldom been considered as a source of live food in aquaculture, partly due to the convenience and the developed techniques associated with the production and use of the dormant cysts. In many countries in Africa, however, hatchery managers do not have access to a reliable supply of affordable good quality cysts. In this study, we therefore demonstrated the potential of a system designed for the continuous ovoviviparous production of nauplii at low salinity, using Great Salt Lake Artemia franciscana and micronized agricultural material as feed. The suitability of the produced nauplii was tested by feeding them directly to Clarias gariepinus larvae in comparison with oviparous nauplii and decapsulated cysts. Higher survival (100%), better protein efficiency ratio (2.6 ± 0.1) and food conversion ratio (1.0 ± 0.1) was observed in larvae fed with the ovoviviparous nauplii (p < 0.05). Overall, we conclude that the ovoviviparous nauplii could serve as an alternative live food for larval fish. If optimized, the system could be validated for integration in hatcheries.