High yield and rapid growth of Neoparamoeba pemaquidensis in co-culture with a rainbow trout gill-derived cell line RTgill-W1

Neoparamoeba pemaquidensis is an ubiquitous amphizoic marine protozoan and has been implicated as the causative agent for several diseases in marine organisms, most notably amoebic gill disease (AGD) in Atlantic salmon. Despite several reports on the pathology of AGD, relatively little is known about the protozoan and its relationship to host cells. In this study, an in vitro approach using monolayers of a rainbow trout gill cell line (RTgill-W1, ATCC CRL-2523) was used to rapidly grow large numbers of N. pemaquidensis (ATCC 50172) and investigate cell-pathogen interactions. Established cell lines derived from other tissues of rainbow trout and other fish species were also evaluated for amoeba growth support. The amoebae showed preference and highest yield when grown with RTgill-W1 over nine other tested fish cell lines. Amoeba yields could reach as high as 5 x 10(5) cells mL(-1) within 3 days of growth on the gill cell monolayers. The amoebae caused visible focal lesions in RTgill-W1 monolayers within 24 h of exposure and rapidly proliferated and spread with cytopathic effects destroying the neighbouring pavement-like cells within 48-72 h after initial exposure in media above 700 mOsm kg(-1). Disruption of the integrity of the gill cell monolayers could be noted within 30 min of exposure to the amoeba suspensions by changes in transepithelial resistance (TER) compared with control cell monolayers maintained in the exposure media. This was significantly different by 2 h (P < 0.05) compared with control cells and remained significantly different (P < 0.01) for the remaining 72 h that the TER was monitored. The RTgill-W1 cell line is thus a convenient model for growing N. pemaquidensis and for studying host-pathogen interactions in AGD.     

Stock structure of blue threadfin Eleutheronema tetradactylum across northern Australia as inferred from stable isotopes in sagittal otolith carbonate

Abstract Stable isotopes of δ¹⁸O and δ¹³C in sagittal otolith carbonates were used to determine the stock structure of the polynemid Eleutheronema tetradactylum (Shaw) across tropical northern Australia, where this species is an important component of inshore commercial and recreational fisheries. Stable isotopes from the sagittal otolith carbonate of 470 fish from 11 discrete locations across western, northern and eastern Australia were sampled between 2007 and 2009. Analysis of these stable isotopes revealed different location‐specific signatures, indicating strong population subdivision. The significant differences in the isotopic signatures of E. tetradactylum demonstrated that there is unlikely to be substantial movement of fish between these locations. The spatial separation of these populations indicates a complex fine spatial scale stock structure across northern Australia, with at least 11 stocks or management units present. The population subdivision of E. tetradactylum was evident along expansive stretches of open beach systems and within coastal embayments with no physical barriers such as headlands. These results indicate that optimal fisheries management will require a review of the current spatial arrangements, particularly the potential for localised depletion of stocks on small spatial scales.     

Influence of dietary fat level and whole-body adiposity on voluntary energy intake by juvenile rainbow trout Oncorhynchus mykiss (Walbaum) under self-feeding conditions

The influence of dietary fat level and whole‐body adiposity on voluntary energy intake of juvenile rainbow trout Oncorhynchus mykiss (Walbaum) was examined using self‐feeders. Groups of lean fish [crude fat (CF) = 7%] and fat fish (CF = 11%), pretreated with a commercial diet with or without supplemental pollock oil, were self‐fed one of three fat level diets (CF = 8%, 13.5% and 19%) for 48 days at 17 °C. Final body weight (BW) and total digestible energy (DE) intake (kJ per fish) were positively affected by the initial BW. Relative to the initial BW, however, fat fish consumed less DE than lean fish. Although the effect of dietary fat level was not significant, percentage weight gain and daily DE intake per BW (kJ kg^?1 BW day^?1) of fat fish were significantly lower than those of lean fish (ancova with initial BW as a covariate, P < 0.05). Energy digestibility, feed efficiency and protein retention were improved with the dietary fat level; however, there was no difference resulting from body fat level. The whole‐body fat levels at the end of the experiment increased with the dietary fat level. Between groups self‐fed the same diet, fat levels of the initially fat fish were still higher than those of the lean fish. The results of the present medium‐term study suggest that rainbow trout adjust DE intake from diets with fat levels ranging from 8% to 19%. Although body fat level affects neither energy digestibility nor protein utilization, a high body fat level may reduce DE intake and consequently depress growth.     

Effect of prebiotic xylooligosaccharides on growth performances and digestive enzyme activities of allogynogenetic crucian carp (<Emphasis Type="Italic">Carassius auratus gibelio</Emphasis>)

The effect of prebiotic xylooligosaccharides (XOS) on the growth performance and digestive enzyme activities of the allogynogenetic crucian carp, Carassius auratus gibelio, was investigated. XOS was added to fish basal semi-purified diets at three concentrations by dry feed weight: diet 1, 50 mg kg⁻¹; diet 2, 100 mg kg⁻¹; diet 3, 200 mg kg⁻¹, respectively. Twelve aquaria (n = 20) with three replicates for each treatment group (diets 1–3) and control treated without XOS were used. Weights of all collected carp from each aquarium were determined at the initial phase and at the end of the experiment, and the carp survival was also determined by counting the individuals in each aquarium. After 45 days, there were significant differences (P < 0.05) in the relative gain rate (RGR), and daily weight gain (DWG) of diets 1–3 were compared with the control. However, the survival rate was not affected (P > 0.05) by the dietary treatments. For enzymatic analysis, dissection produced a crude mixture of intestine and hepatopancreas of each segment to measure. The protease activity in the intestine and hepatopancreas content of fish in diet 2 (487.37 ± 20.58 U g⁻¹ and 20.52 ± 1.93 U g⁻¹) were significantly different (P < 0.05) from that in the control (428.13 ± 23.26 U g⁻¹ and 12.81 ± 1.52 U g⁻¹) and diet 3 (428.00 ± 23.78 U g⁻¹ and 14.04 ± 1.59 U g⁻¹). Amylase activity in the intestine was significantly higher for diet 2 compared to diet 1 and the control. As for amylase in the hepatopancreas, assays showed higher activity in diet 2 (P < 0.05) compared to the rest.     

Artificially induced tetraploid masu salmon have the ability to form primordial germ cells

Diploid gametes generated with tetraploid animals are a stepping stone to improving chromosome manipulation techniques. However, artificially induced tetraploid individuals generally die soon after hatching. Diploid gametes could be induced by in vivo cultures of tetraploid primordial germ cells (PGCs) through germ-line chimera. In the present study, characteristics of PGCs were studied in inviable tetraploid masu salmon, Oncorhynchus masou. Histological observation of tetraploid embryos revealed that the same or smaller numbers of PGCs were observed and they migrate into the genital ridges as did diploid PGCs during gonadogenesis. By whole-mount in situ hybridization using vasa messenger RNA (mRNA), 4–35 vasa-positive signals were detected in a pair of genital ridges of tetraploids. By cytological observation of genital ridge cell suspensions, several large round cells were observed, some of which extended pseudopodia. They also contained large nuclei and round granules in their cytoplasm, characteristics of PGCs. As the results suggest that inviable artificial tetraploids have PGCs, we expect to achieve diploid gamete production through surrogate propagation and tetraploid fish production.     

Effect of cryoprotectants, extenders and freezing rates on the fertilization rate of frozen striped catfish, Pangasius hypophthalmus (Sauvage), sperm

The effects of four cryoprotectants (methanol, MeOH; dimethyl sulphoxide, DMSO; dimethyl acetamide, DMA; and ethylene glycol, EG), three extenders (calcium‐free Hanks' balanced salt solution, C‐F HBSS, Hanks' balanced salt solution, HBSS and sodium chloride, NaCl) and two different freezing procedures (one‐ and two‐step) on the cryopreservation of striped catfish (Pangasius hypophthalmus (Sauvage)) sperm were investigated. Sperm were frozen using a controlled‐rate freezer in 250 μL straws and stored for 2 weeks in a liquid nitrogen (LN₂) container. They were then airthawed at room temperature, and fertilization, motility and viability were assessed. The highest fertilization rate of 41% (81% of control) was achieved with the combination of 12% DMSO and 0.9% NaCl using a one‐step freezing procedure (10°C min^?1). Also, DMA resulted in a higher fertilization rate (30% or 51% of the control) than MeOH (18% or 38% of the control) or EG (8% or 12% of the control). In addition, the three extenders used did not affect fertilization rates after cryopreservation with each cryoprotectant. There were no significant differences among the three cryoprotectant concentrations and between the one‐ and two‐step freezing procedures. However, fertilization rates of cryopreserved sperm were significantly lower than the controls (P<0.05). The results of this study indicate that high fertilization rates of striped catfish eggs can be achieved using cryopreserved sperm when frozen at 10°C min^?1 in DMSO or DMA with either 0.9% NaCl or C‐F HBSS.     

Nutritional evaluation of decapsulated cysts of fairy shrimp (Streptocephalus dichotomus) for ornamental fish larval rearing

The suitability of decapsulated cysts of the fairy shrimp, Streptocephalus dichotomus as a sole diet was evaluated for the ornamental angelfish Pterophylum scalare larvae. Brine shrimp, Artemia sp., and microworms, Panagrellus redivivus, were used as comparison foods. The results indicate an appreciable weight gain of 15.22±1.69 mg and a growth (length) of 0.876±0.03 cm in experimental fish fed decapsulated fairy shrimp cysts compared with the growth and weight of fish fed other foods. The influence of the experimental diets was further reflected in the composition of fatty acid and amino‐acid profiles of the experimental fish. Angelfish larvae readily consumed the decapsulated cysts and utilized them efficiently during the early days of exogenous feeding.     

Apparent digestibility coefficients of nutrients in brewer's and rendered animal by-products for rainbow trout (Oncorhynchus mykiss (Walbaum))

Two experiments were conducted to measure the apparent digestibility coefficients (ADCs) of nutrients, dry matter, and energy in by‐products of the brewing industry and in selected animal protein ingredients for rainbow trout. In experiment 1, 500 rainbow trout (average body weight 170.8±5.5 g) were stocked in ten 140‐L digestibility tanks with 50 fish per tank and two tanks per diet. Yttrium oxide was used as an inert marker in the diets. The high fibre content of brewer's dried grains (BDG) affected the ADCs of dry matter but not of protein or amino acids. Brewer's dried yeast had a higher protein content than BDG, but ADC values for protein and amino acids were significantly lower. The ADCs of phosphorus were similar among brewer's dried products. The BDG high‐protein, fraction had marginally higher ADC values for dry matter, protein and amino acids than regular BDG. Conditions in experiment 2 were similar to those in experiment 1. ADC values for spray‐dried porcine plasma were over 98% for dry matter, crude protein, crude fat, and gross energy. ADC values for spent hen meal were higher than those of poultry by‐product meal or feather meal, with the exception of gross energy. However, spent hen meal was unpalatable. The ADCs in these ingredients were variable, and this variability must be taken into account when these ingredients are formulated into feeds for fish.