Nematode parasites infecting the starry batfish Halieutaea stellata (Vahl) (Lophiiformes: Ogcocephalidae) from the East and South China Sea

The starry batfish Halieutaea stellata (Vahl) is a small, benthic fish found in Indo‐West Pacific Oceans. However, our present knowledge of the helminth parasites of this fish is still fragmentary. In this study, a total of 29 fish collected from the East and South China Sea were examined to determine the prevalence, intensity and species composition of helminth parasites in H. stellata. Using morphological and molecular approaches, four species of nematodes were found parasitic in this fish host, including the adults and fourth‐stage larvae of Raphidascaroides nipponensis Yamaguti 1941; adults and third‐stage larvae of Raphidascaris lophii (Wu 1949), third‐ and fourth‐stage larvae of Hysterothylacium larval type IV‐A of Shamsi, Gasser & Beveridge 2013 and third‐stage larvae of Hysterothylacium amoyense (Hsü 1993). Halieutaea stellata represents a new host record for the three last‐named nematodes. Raphidascaroides nipponensis with the highest prevalence (82.5%) and intensity (mean = 13.5) of infection was considered as the dominant parasite species in H. stellata. The detailed morphology of the different developmental stages of the four nematode species was studied using light and scanning electron microscopy. All nematode species were also genetically characterized by sequencing and analysing the internal transcribed spacer (ITS) of the ribosomal DNA. This study provides further data on the occurrence of nematode parasites in H. stellata and also contributes to facilitate an accurate and rapid diagnosis of the infection by these little‐known nematodes.     

The hidden companion of non‐native fishes in north‐east Atlantic waters

A tropicalization phenomenon of ichthyofauna has been described in the last decades in Galicia (north‐eastern Atlantic), with increasing reports of tropical and subtropical fishes appearing northward this distribution range. A search for parasites was carried out in the digestive tract of two specimens first captured in Galician waters: the prickly puffer Ephippion guttifer (Tetraodontidae) and the African stripped grunt Parapristipoma octolineatum (Haemulidae). Examination of E. guttifer showed high intensity of nematodes, from three different genera: Cucullanus (Cucullanidae), Hysterothylacium (Raphidascaridae) and Anisakis (Anisakidae), with demonstrated pathogenicity to humans. Molecular identification allowed the identification of Anisakis pegreffii, already described in the area, and first reports for European waters of Cucullanus dodsworthi, Hysterothylacium reliquens and a new Hysterothylacium sp. P. octolineatum showed a far lower level of parasitization, with two Hysterothylacium larvae, genetically identified as Hysterothylacium deardorffoverstreetorum, also its first report in the eastern Atlantic. Thus, possible ecological impact of the occurrence of two non‐native individual fishes in a new area could be remarkably higher if we see this issue through the lens of the parasitological perspective, as far as only two individual fish can harbour more of one hundred nematode parasites belonging to different species, most of them also new species for that area.     

Reproduction,larviculture and early development of the Bluebanded goby,Lythrypnus dalli,an emerging model organism for studies in evolutionary developmental biology and sexual plasticity

The Bluebanded goby, Lythrypnus dalli, is a popular ornamental aquarium species and a key organism for the study of several fundamental biological questions, most notably reversible sex change in adults. To maximize the tractability of this species as an emerging model system, it is essential to have an optimized propagation system and a detailed developmental staging scheme. One limitation to the larviculture of L. dalli is the relatively small size of the larvae, which makes the transition from yolk to feeding challenging. We developed a protocol and successfully reared three generations of L. dalli in the laboratory. The protocol contains several key innovations for the rearing of diminutive fish larvae, including tank design and co‐culturing of microalgae (Isochrysis galbana) with copepods (Parvocalanus sp.) in the larval rearing tanks. In addition, we describe the embryonic and larval development of L. dalli under controlled conditions and in comparison with the model organism Danio rerio. We found that at 21°C L. dalli larvae hatch in 4 days, reach flexion in 18–25 days and are sexually mature by 3 months. Overall, the embryonic development of L. dalli is remarkably similar to D. rerio with several striking differences, including the position and shape of the blastomere, size of the neuromasts and corresponding cupula, and relative timing of pigmentation and brain subdivision. The ability to rear this species in captivity is a valuable tool that could be utilized for a variety of similarly diminutive species and to address a greater breadth and depth of biological questions.     

Understanding imperfect detection in a San Francisco Estuary long‐term larval and juvenile fish monitoring programme

Imperfect detection can present a significant challenge when monitoring for a rare and imperilled species. Here, a long‐term larval and early‐juvenile fish monitoring programme in the upper San Francisco Estuary was examined to evaluate its overall reliability in detecting various fish species, including the imperilled delta smelt, Hypomesus transpacificus McAllister, for which the programme was designed. Using occupancy modelling, detection probability of species with pelagic larval or juvenile life stages was found to be generally high (≥.95) based on the current sampling effort of three larval net tows per site. However, detection probability can vary considerably from year to year depending on the species’ level of larval production. Water temperature and turbidity were identified as important predictors of occurrence for young‐of‐year delta smelt, longfin smelt Spirinchus thaleichthys (Ayres) and striped bass Morone saxatilis (Walbaum), and there was evidence for fish size selectivity by the sampling gear in all three species. These results highlight the need to consider adaptively managing detection probability by increasing sampling effort in years when young‐of‐year delta smelt abundance is expected to be low, especially when information on the species’ occurrence at a particular region is critical.     

Does upwelling intensity determine larval fish habitats in upwelling ecosystems? The case of Senegal and Mauritania

European sardine (Sardina pilchardus) and round sardinella (Sardinella aurita) comprise two‐thirds of total landings of small pelagic fishes in the Canary Current Eastern Boundary Ecosystem (CCEBE). Their spawning habitat is the continental shelf where upwelling is responsible for high productivity. While upwelling intensity is predicted to change through ocean warming, the effects of upwelling intensity on larval fish habitat expansion is not well understood. Larval habitat characteristics of both species were investigated during different upwelling intensity regimes. Three surveys were carried out to sample fish larvae during cold (permanent upwelling) and warm (low upwelling) seasons along the southern coastal upwelling area of the CCEBE (13°–22.5°N). Sardina pilchardus larvae were observed in areas of strong upwelling during both seasons. Larval habitat expansion was restricted from 22.5°N to 17.5°N during cold seasons and to 22.5°N during the warm season. Sardinella aurita larvae were observed from 13°N to 15°N during cold seasons and 16–21°N in the warm season under low upwelling conditions. Generalized additive models predicted upwelling intensity driven larval fish abundance patterns. Observations and modeling revealed species‐specific spawning times and locations, that resulted in a niche partitioning allowing species' co‐existence. Alterations in upwelling intensity may have drastic effects on the spawning behavior, larval survival, and probably recruitment success of a species. The results enable insights into the spawning behavior of major small pelagic fish species in the CCEBE. Understanding biological responses to physical variability are essential in managing marine resources under changing climate conditions.     

Entrainment and advection of larval sardine,Sardinops sagax,by the East Australian Current and retention in the western Tasman Front

The poleward flowing East Australian Current (EAC) drives sporadic upwelling, entrains coastal water and forms the western Tasman Front (wTF), creating a mosaic of water types and larval transport routes along south eastern Australia. The spatial distribution, otolith chemistry and growth rates of larval sardine (Sardinops sagax) were examined to infer spawning location and larval transport. A gradient of increasing larval size from north to south along the shelf was not detected but was evident between the shelf and offshore in the wTF. Here larvae were larger and older. Based on the occurrence of newly hatched larvae, spawning by S. sagax between southern Queensland and mid New South Wales (NSW) was more extensive than previously reported. The otolith chemistry from two wTF larval size classes differed, implying different origins. The otolith chemistry of wTF post‐flexion larvae was similar to larvae from northern NSW, whereas wTF flexion larvae were similar to larvae observed nearby from mid‐NSW. Two possible larval transport routes, direct and indirect, are inferred from otolith chemistry, current velocities and a previously published particle tracking study. Either larvae from northern NSW were advected south and entrained with younger larvae directly into the wTF, or larvae from a range of shelf regions were advected around the southern edge of an anticyclonic eddy, to join younger larvae directly entrained into the wTF. Based on the co‐occurrence of larval ages and sizes in the wTF and their advection routes, the wTF appears to be an important larval retention zone.     

The settlement and reproductive success of Lepeophtheirus salmonis (Krøyer 1837; Copepoda: Caligidae) on atypical hosts

The reproductive success of Lepeophtheirus salmonis settled on host and non‐host fish has been compared. Triplicate single species tanks of Atlantic salmon, marine three‐spined sticklebacks, saithe and Atlantic cod were exposed to 10 adult female L. salmonis per tank (n=30 lice per species). Adult female L. salmonis settlement and egg string production occurred only on salmon and cod, with no egg production occurring on saithe and three‐spined sticklebacks. The number of eggs in egg strings, hatching success of eggs and the survival of all larval stages to the copepodid stage were severely affected by the species of fish on which female L. salmonis had settled. L. salmonis settled on cod produced significantly fewer eggs, lower hatching rates and lower survival rates of larvae than females on Atlantic salmon. The production of egg strings by L. salmonis females infecting cod, which successfully hatch and moult through to the infective copepodid stage, albeit in small numbers, is discussed in terms of the implications to aquaculture and salmon and cod farming scenarios.     

Comparative assessment of Vibrio virulence in marine fish larvae

Vibrionaceae infections are a major obstacle for marine larviculture; however, little is known about virulence differences of Vibrio strains. The virulence of Vibrio strains, mostly isolated from vibriosis outbreaks in farmed fish, was tested in larval challenge trials with cod (Gadus morhua), turbot (Scophthalmus maximus) and halibut (Hippoglossus hippoglossus) using a multiwell dish assays with single‐egg/larvae cultures. The strains differed significantly in virulence as some caused a high mortality of larva reaching 100% mortality after a few days, while others had no or only marginal effects on survival. Some Vibrio strains were pathogenic in all of the larva species, while some caused disease only in one of the species. Twenty‐nine of the Vibrio anguillarum strains increased the mortality of larvae from at least one fish species; however, pathogenicity of the strains differed markedly. Other Vibrio species had no or less pronounced effects on larval mortalities. Iron uptake has been related to V. anguillarum virulence; however, the presence or absence of the plasmid pJM1 encoding anguibactin did not correlate with virulence. The genomes of V. anguillarum were compared (D. Castillo, P.W. D'Alvise, M. Middelboe & L. Gram, unpublished data) and most of the high‐virulent strains had acquired virulence genes from other pathogenic Vibrio.     

Effects of docosahexaenoic acid on growth,survival and swim bladder inflation of larval amberjack ( Seriola dumerili, Risso)

The effect of docosahexaenoic acid (DHA) on the growth performance, survival and swim bladder inflation of larval Seriola dumerili during the rotifer feeding period was investigated in two feeding experiments. Amberjack larvae at 3 day post hatching were fed rotifers enriched with (1) freshwater C hlorella (Chlo), (2) a mixture (2:1, v/v) of Chlo and DHA‐enriched C hlorella (DHA‐Chlo), (3) DHA‐Chlo and (4) DHA‐Chlo and commercial DHA emulsion, in triplicate for 7 days. The average DHA contents of the rotifers were 0.0, 0.4, 1.0 and 1.9 mg g^?1 DM respectively. The survival rate was improved by the enrichment of rotifers with DHA‐Chlo alone, and DHA‐Chlo and emulsion. Growth and swim bladder inflation of fish fed rotifers enriched with DHA‐Chlo were significantly (P < 0.05) improved, however, with increased levels of DHA further improvement was not found. DHA content in the larval whole body proportionally increased with the DHA level in the rotifers. These results suggest that DHA enrichment of rotifers is effective to improve the growth, survival rate and swim bladder inflation of amberjack larvae. The DHA requirement of amberjack larvae is estimated to be 1.5 mg g^?1 on a dry matter basis of rotifers.     

Optimizing stocking density and microalgae ration improves the growth potential of tropical black‐lip oyster,Saccostrea echinata,larvae

The combined effects of stocking density and microalgae ration on survival and size of Saccostrea echinata larvae were studied in two‐factor experiments for the major developmental stages: D‐veliger (1‐day posthatch [dph], Experiment 1), umbonate (12 dph, Experiment 2), and eyed (19 dph, Experiment 3) larvae. Larvae were stocked into replicate sets of four 10‐L aquaria with ambient 1‐μm filtered sea water (28 ± 1.5°C and 36 ppt) and cultured for four days at densities of 0.5, 2, 5, 7, or 10 larvae/mL and provided with microalgae rations at each of five densities (cells larvae^?1 day^?1); 0, 1, 3, 5, or 8 × 10³ (D‐veliger larvae, Experiment 1); 0, 5, 12, 18, or 25 × 10³ (umbonate larvae, Experiment 2); and 0, 15, 30, 40, or 60 × 10³ (eyed larvae, Experiment 3). Microalgae rations for each larval life stage were selected on the basis of increasing food requirement with larval size and comprised a 2:1:1 mixture of Chaetoceros calcitrans, Tisochrysis lutea, and Pavlova spp., calculated on an equal dry‐weight basis. Contour plots were generated from larval survival and larval size (dorso‐ventral measurement [DVM]) data to determine optimal culture conditions. Larvae showed high survival (54–100%) over a wide range of both treatment parameters across all life stages, confirming broad tolerance limits for this species. The interaction effects of larval stocking density and microalgae ration on larval size were significant (p < 0.001) across all life stages. Results indicate that maximum larval size (DVM) is achieved when S. echinata are cultured at: 6–8 larvae/mL and fed 5–6 × 10³ cells larvae^?1 day^?1 for D‐veligers (mean DVM >80 μm), at 2–8 larvae/mL and fed 11–25 × 10³ cells larvae^?1 day^?1 for umbonate larvae (mean DVM > 190 μm), and at 1–4 larvae/mL and fed 15–40 × 10³ cells larvae^?1 day^?1 for eyed larvae (mean DVM >230 μm). Results will help refine current hatchery methods for S. echinata supporting further development toward commercial aquaculture production of this species.     

In vitro cultivation model for Heterosporis saurida (Microsporidia) isolated from lizardfish,Saurida undosquamis (Richardson)

Heterosporis saurida is a microsporidian that infects lizardfish, Saurida undosquamis (Richardson, 1848), in the Arabian Sea. Spores were isolated from infected lizardfish and used to infect derived fish cell lines: common carp brain (CCB), epithelioma papulosum cyprinid (EPC), fathead minnow epithelial (FHM), rainbow trout gonad (RTG), bluegill fry (BF‐2) and chinook salmon embryo (CHSE). Non‐fish cell lines were also tested that include: insect (SF‐9), rabbit (RK‐13) and African green monkey (Vero E6). No growth of H. saurida was observed in any fish cell line, SF‐9 or Vero E6 cell lines. H. saurida spores grew only in RK‐13 cell line and were detected by immunofluorescence. Developmental stages of H. saurida were seen in RK‐13 cells by light and transmission electron microscopy, and species identification was confirmed by sequencing. This study demonstrated that H. saurida was able to proliferate in the mammalian RK‐13 cell line, which thus represents an in vitro model for conducting molecular genetics and cell–pathogen interaction studies of Heterosporis.     

Assessment of dietary lipid sources in tropical gar,Atractosteus tropicus larvae: Growth parameters and intermediary lipogenic gene expression

Five experimental diets containing different lipid sources, fish oil (D1), soybean lecithin (D2), corn oil (D3), canola oil (D4) and olive oil (D5), were evaluated in Atractosteus tropicus larvae for the relative gene expression of the enzymes fatty acid synthase (fas), acetyl‐CoA carboxylase 1 (acc1) and carnitine palmitoyltransferase 1C (cpt1c), in addition to their effects on larval growth, survival and cannibalism during a 30‐day feeding trial. Higher growth and survival were obtained in treatments D1 and D2, and lower performance in diets D3, D4 and D5. The highest levels of expression of fas and acc1 occurred in larvae fed with D1, which contained high amounts of n‐3 long‐chain polyunsaturated fatty acids (LC‐PUFA), mainly DHA and EPA FA are regulators of lipogenesis. The higher cpt1c expression in plant‐based diets is attributed to the fact that these diets are rich in α‐linolenic acid (ALA) and low DHA, EPA and ARA levels that favour ß‐oxidation. In conclusion, the diets with fish oil (D1) and soybean lecithin (D2) were the best treatments for larval growth, survival and cannibalism and thus appear to meet both lipid and energy requirements of A. tropicus larvae, meanwhile the use of vegetable oils influences the expression of intermediary lipogenic genes.     

In Vitro Protein Digestibility of Dietary Ingredients Throughout Ontogeny of California Halibut,Paralichthys californicus,Larvae

Digestive capacity varies substantially in marine fish larvae during development, and the formulation of larval diets does not take this into account. The objectives of this study were to assess in vitro protein digestibility of dietary ingredients throughout the ontogeny of digestive system in California halibut, Paralichthys californicus, larvae so as to identify adequate protein sources for each stage of development. Protein digestibility of potential sources was evaluated using the pH‐STAT technique. Digestive enzymes utilized for the in vitro assays were obtained from homogenates of whole bodies (9, 15, and 26 days post hatch [dph]) and dissected guts of larvae (39 and 51 dph) sampled at different stages of development. Significant differences in degree of protein hydrolysis (DH) were found among developmental stages for some ingredients. Meals elaborated with rotifers and Artemia showed the highest DH with a tendency to decrease as larval development progressed. Digestibility of casein was poor early in development and increased as it progressed. In contrast, soybean and krill meals were poorly digested throughout development, which may be attributed to differences in the nature or quality of the ingredient. Results from this study highlight the importance of evaluating the digestibility of protein sources through larval development to formulate successful, stage‐specific weaning diets.     

Use of Ox‐Aquaculture© for disinfection of live prey and meagre larvae,Argyrosomus regius (Asso, 1801)

Live prey used for marine larval fish (rotifers and Artemia) as well as intensive larval rearing conditions are susceptible to the proliferation of bacteria that are the cause for reduced growth and larval mortality. Hydrogen peroxide has been recently proved a good disinfectant in aquaculture, either for eggs, larvae or live prey. In this study the effects of a hydrogen peroxide‐based product, Ox‐Aquaculture^©, on live prey (rotifers and Artemia) and meagre larvae bacterial load, composition and final status have been tested. A 34.6% reduction of total heterotrophic bacteria and 59.7% of Vibrionaceae were obtained when rotifers were exposed for 15 min to 40 mg L^?1 of the product. A 34.3% reduction of total heterotrophic bacteria and 37.7% of Vibrionaceae were obtained when Artemia were exposed for 5 min to 8000 mg L^?1 of the product. More than 95% reduction of total heterotrophic bacteria and 75% of Vibrionaceae were obtained when meagre larvae were exposed for 1 h to 20 mg L^?1 of the product. Furthermore, disinfection of enriched live prey with the product did not change the fatty acid composition and survival of the live prey and improved final larval survival.     

Embryonic and larval developments of brackish water catfish,Mystus gulio (Hamilton and Buchanan, 1822) induced with human chorionic gonadotropin and consequent larval rearing

Mystus gulio, the long whiskers catfish, is a popular food fish and potential candidate species for aquaculture in Sundarban area of India and Bangladesh. Recently, catch of this species has declined due to overfishing and various ecological changes. In the present study, mature fish was induced to spawn in captivity through intramuscular injection of human chorionic gonadotropin at the doses of 10 IU/g to female and 5 IU/g to male. Photomicrographs of all developmental stages of live embryo and larvae were documented with the aid of a light microscope. Results demonstrated that morula, blastula, gastrula and neurula and organogenesis ended at 1:30‐, 3:00‐, 5:30‐, 7:30‐ and 17:15‐hour post‐spawning (hps) respectively. Heartbeat and muscular contraction of embryo commenced at 8:30 and 11.15 hps respectively. Hatching of embryo started after an incubation period of 17:30 hour at an ambient temperature of 29 ± 1°C. The newly hatched larvae measured 2.17 ± 0.29 mm in total length with a yolk volume of 0.165 ± 0.03 mm³ started feeding 36 hr after hatching. The present study, on induced breeding and chronological development of M. gulio embryo, will have significant implications on conservation and seed production for aquaculture.     

The enrichment of rotifers (Brachionus plicatilis) and Artemia franciscana with taurine liposomes and their subsequent effects on the larval development of California yellowtail (Seriola lalandi)

Taurine is an essential or conditionally essential nutrient for many species of marine fish, especially during early development. There is growing evidence that marine fish larvae benefit from taurine‐enriched rotifers; however, it is unknown if larvae benefit from taurine‐enriched Artemia. We investigated the effects of taurine‐enriched rotifers (Brachionus plicatilis) and Artemia franciscana on the growth and whole‐body taurine concentrations of California yellowtail (Seriola lalandi; CYT) larvae. The approach used in this study was to encapsulate taurine within microparticles (liposomes), which were then fed to rotifers and Artemia. We found that feeding taurine liposomes to rotifers and Artemia resulted in taurine concentrations in these prey species that were similar to or above those previously reported in copepods. At the end of the rotifer phase, CYT larvae fed taurine‐enriched rotifers showed increased growth (final dry weights; DW) and had higher whole body taurine concentrations when compared to larvae fed unenriched rotifers. At the end of the Artemia phase, CYT whole body taurine concentrations varied among dietary treatments. Larval lengths and DWs were not significantly different among treatments at the end of the Artemia phase, suggesting that the taurine concentrations of unenriched Artemia were sufficient to support the growth of CYT larvae.     

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).