Verification of ploidy level in sturgeon larvae

Chromosome preparations and assay of the microsatellite locus Afu‐68 were used to determine ploidy in Siberian sturgeon (Acipenser baeri Brandt) and F₁ hybrids of Siberian sturgeon and Russian sturgeon (Acipenser gueldenstaedti Brandt). The chromosome number and microsatellite locus Afu‐68 were compared and these analyses were used for identification of ‘haploid’, ‘diploid’ and ‘triploid’ progeny of the studied cross of A. baeri× (A. baeri×A. gueldenstaedti).     

Triploidization in rohu × mrigal hybrid and comparison of growth performance of triploid hybrid

The present study was aimed at the identification of treatment optima to induce triploidy in ‘Labeo rohita (rohu) × Cirrhinus cirrhosus (mrigal)’ hybrid using heat shock treatment. The eggs were exposed at four different temperature regimes viz., 38, 39, 40 and 41°C for 1–3 min, applied 3–5 min after fertilization. After 4 min of fertilization, heat shock treatments for 1 and 1.5 min durations were found the best inducing triploidy up to 100% and 96% respectively. Survival rates upto yolk sac absorption were found to be 73% and 71% in rohu and mrigal, 68% and 67% in the reciprocal diploid hybrids and 61% and 60% in the reciprocal triploid hybrids (RTH). Triploidy was confirmed by chromosome counting that revealed the diploid chromosome number of rohu and mrigal at 2n = 50 and in their triploid hybrid chromosome number was found to be 3n = 75. Growth rate of the RTH showed a significant difference (P < 0.05) from the single species and the diploid hybrids. Triploids also showed higher survival rate over the diploids.     

Inter‐specific hybridization between Black Sea trout (Salmo labrax Pallas, 1814) and rainbow trout (Oncorhynchus mykiss Walbaum, 1792)

Thermal shock‐induced triploid and unshocked control hybrids between rainbow trout (Oncorhynchus mykiss) and Black Sea trout (Salmo labrax) and their parental species were produced under hatchery condition by using heat shocks. Triploidization reduced eyed egg rate and alevin yield in all groups. Low survival rate was observed in both shock‐induced triploid hybrid and non‐shock‐induced control hybrids. Although hybrids demonstrated low body weight during the first feeding stage, they reached higher body weight by day 200 when compared with Black Sea trout and rainbow trout. A higher specific growth rate was calculated as 3.60 in the triploid hybrid groups, 1.41 in the triploid Black Sea trout groups and 2.27 in the triploid rainbow trout groups between days 110 and 200. A lower condition factor was determined in the hybrid than in the diploid parental species. A negative value of mid‐parent heterosis (MPH) performance was deected for condition factor, and a favourable MPH was detected for specific growth rate and weight in hybrids.     

Characterization of triploid hybrid groupers from interspecies hybridization (Epinephelus coioides ♀ × Epinephelus lanceolatus ♂)

Interspecies hybridization is widely used in aquaculture as a beneficial strategy. Diploid and triploid hybrids have been detected from the interspecies hybridization of Epinephelus coioides ♀ × Epinephelus lanceolatus ♂. This is the first report of triploidization through hybridization in grouper. Confirmation has been obtained through flow cytometry, karyotyping and erythrocyte nuclei measurement. The chromosome numbers of E. coioides, E. lanceolatus, diploid hybrid grouper are 48 and triploid hybrid grouper are 72. Measurements of erythrocyte nuclei indicate that triploid fish have a larger nuclear surface than the diploid groupers, and the average ratio of triploid to diploid surface area is 1.59. During the first 1.5 years, triploid hybrid groupers grow faster than diploid hybrid groupers or either parent species. The average growth rate of triploid hybrids is 1.61 times greater than that of diploid hybrids at 6 months of age and 1.43 times greater at 18 months of age. The triploid hybrid groupers are inferior in gonadal development, with no primary‐growth‐stage oocytes appearing in the gonads at 18 months of age. Morphological studies indicate that triploid hybrid groupers have distinctive differences in snout length, eye diameter, body trunk shape, and tail shape development compared with diploid hybrid groupers. Triploid hybrid groupers have an advantage in growth ability, and artificial breeding of triploid groupers might be of great potential use in the grouper aquaculture industry.     

Reproductive ability of triploid ornamental (koi) carp (Cyprinus carpio L.) × goldfish (Carassius auratus L.) hybrids and characteristics of their offspring

The purpose of this study was to investigate reproductive ability of backcross triploid koi (Cyprinus carpio L.) × goldfish (Carassius auratus L.) hybrids. These triploids have been obtained by crossing of F₁ hybrid females producing diploid eggs with males of parental species. Triploid hybrid females, when crossed with goldfish or koi males, produced mostly aneuploid fish with ploidy range from approximately 2.2n–3.2n with a mean value 2.5n; some fish in crosses of triploid females with koi males were tetraploid (4.0n). Since analysed fish had in their genomes one haploid set from parental males, the data indicate that triploid hybrid females mostly produced aneuploid eggs with ploidy range from approximately 1.2n–2.2n and a mean ploidy around 1.5n while some eggs were triploid. Triploid hybrid males were completely sterile and have not released any sperm after hormonal injection. Despite their low viability, some aneuploid fish obtained from triploid hybrid females were raised in indoor recirculating systems until the age of 2 years and their reproductive ability has been evaluated. One aneuploid female with ploidy 2.1n produced larvae with ploidy range from 2.9n to 3.4n with a mean ploidy of 3.1n when crossed with a koi male; about 60% of obtained larvae had ploidy from 3.0n to 3.2n. These data indicate that this female produced mostly eggs with unreduced ploidy level.     

Saltwater performance of triploid Atlantic salmon Salmo salar L. brown trout Salmo trutta L. hybrids

Survival and growth in a saltwater net-pen of sexually immature triploid Atlantic salmon Salmo salar L. brown trout Salmo trutta L. hybrids was comparable to that of immature diploid Atlantic salmon. Following 17 months of freshwater rearing, the experimental fish were individually tagged and transferred to a saltwater net-pen where they were raised communally for 376 days. Initial and final average weights were 158 and 760 g per fish for the diploid Atlantic salmon and 209 and 1010 g per fish for the triploid hybrids; weights for the hybrids were significantly larger in both cases (α= 0.05). Survival from transfer to harvest was 43% for the Atlantic salmon and 48% for the hybrids. Average specific growth rate of fish which survived to harvest was 0.42% day^-1 for Atlantic salmon and 0.41% day^-1 for hybrids; these values were not significantly different. No significant differences were observed in average condition factor and dress-out percentage between crosses. Average gonadal weights and gonadosomatic indices were not significantly different for male diploid Atlantic salmon, female triploid hybrids and male triploid hybrids, but were significantly greater for female Atlantic salmon.     

Growth performance, morphometric traits and gonad development of induced reciprocal diploid and triploid hybrids between the mud loach (Misgurnus mizolepis Günther) and cyprinid loach (Misgurnus anguillicaudatus Cantor)

Two species of loach, the mud loach (Misgurnus mizolepis Günther) and the cyprinid loach (M. anguillicaudatus Cantor), are commercially important in Korea both for food and ceremonial purposes. The mud loach has superior potential for aquaculture in terms of growth, whereas the cyprinid loach has a more desirable body shape and colour. This study was conducted to produce reciprocal diploid and triploid hybrids and to evaluate their potential benefits in culture by examining growth performance, morphometrics and gonad development. Reciprocal diploid and triploid hybridization was performed by artificial insemination without or with the induction of triploidy. The successful formation of diploid or triploid karyogamy was verified by flow cytometric analysis. Body weights of induced reciprocal diploid hybrids were intermediate between those of the two parental species, i.e. hybrids were heavier than cyprinid loach but lighter than mud loach. In contrast, the growth performance of triploid hybrids was similar to that of their corresponding maternal parents. Diploid hybrids showed intermediate morphometric traits between the two species; however, the body proportions of triploid hybrids were more similar to those of the maternal species than the paternal species. Histological analyses indicated that reciprocal diploid hybrids of both sexes were able to reach maturity, as evidenced by the presence of mature oocytes or spermatozoa in the gonad tissues. However, triploid hybrids showed stringent sterility at the gonadic level; the sizes of ovaries and testes were much smaller, and gonad development was abnormal and significantly retarded.     

Reproductive ability of second generation ornamental (koi) carp (Cyprinus carpio L.) × goldfish (Carassius auratus L.) hybrids and characteristics of their offspring

The purpose of this study was to investigate the reproductive ability of second generation (F₂) koi (Cyprinus carpio L.) × goldfish (Carassius auratus L.) hybrids. Only diploid F₂ males and females were fertile and used in crosses. A significant increase was recorded in male fertility in F₂ versus F₁. In contrast with an earlier study in which only one fertile F₁ male was found, about 20% of F₂ males produced sperm. The observed reproductive ability of F₂ hybrids was similar to that demonstrated by the only fertile F₁ male and F₁ females. F₂ males produced diploid spermatozoa and generated triploids when crossed with koi females. All triploid fish in these progenies were males indicating that F₂ males had a sex chromosome constitution of XY. F₂ females produced diploid eggs and generated mostly triploids when crossed with koi males. In progenies obtained by crosses of F₂ males with F₁ and F₂ females, most of the surviving juveniles (63%–100%) were diploid; a minority of juveniles were aneuploid (ploidy ranged from 2.1n to 3.6n). Diploid fish in these progenies were presumably the result of spontaneous androgenesis and gynogenesis, by the same mechanisms observed earlier in progenies obtained by crossing the F₁ male with F₁ females.     

Susceptibility of koi × crucian carp and koi × goldfish hybrids to koi herpesvirus (KHV) and the development of KHV disease (KHVD)

Hybrids of koi, Cyprinus carpio × crucian carp, Carassius carassius and koi × goldfish, Carassius auratus, proved to be susceptible to koi herpesvirus (KHV, syn. CyHV‐3) and developed KHV disease (KHVD). While hybrids of koi × goldfish were partly resistant to mortality following infection by immersion, most koi × crucian carp hybrids died after bath infection. KHV DNA was detected in dead fish but also in all surviving animals by different polymerase chain reactions (PCRs). According to these results, hybrid crossbreeding does not seem to prevent severe losses associated with KHV in terms of inducing KHVD. The present study showed severe losses after a waterborne KHV infection of between 35% and 100% in koi × goldfish and koi × crucian carp hybrids as well as in SPF carp.     

Effect of Stock Density and Ploidy in Jundia,Rhamdia quelen,Larvae Performance

The performance (growth and survival) of diploid and triploid jundia, Rhamdia quelen, was evaluated at six different stocking densities (10, 60, 110, 160, 210, 260 larvae/liter) during 31 days after rearing in an intensive larviculture system. Triploid fish exhibited a significantly higher survival rate than diploids at all stocking densities (27.1 ± 4.3% vs. 12.1 ± 3.3%; P < 0.01). Survival was not affected by stocking density (P > 0.05). Length gain was not affected by either ploidy or stocking density. Diploid fish gained more weight than triploids (P < 0.05), though this difference could result from lower fish densities in diploid treatments resulting from the higher mortality rate of diploid fish. This hypothesis is strengthened by the higher biomass present in triploid treatments (P < 0.01).     

Comparison of freshwater and marine performances of all‐female diploid and triploid Atlantic salmon (Salmo salar L.)

The performance of all‐female diploid (AF2N) and triploid (AF3N) Atlantic salmon were compared in fresh water, under commercial production conditions in 1995 and 1996 year classes. The performance of the 1996 year class was also assessed for 14 months in a commercial sea farm. Freshwater mortality was higher in the triploid groups. The majority of losses occurred in the early stages of egg development and during the first feeding period, when the incidence of non‐feeding fry was consistently higher. In growth studies, although diploid fry were significantly heavier during first feeding there were no significant differences in weight between groups some 8 months after fertilization or in presmolt growth periods from February to April in 1996 and 1997. Smolting rates were high (range 93.5–95.3%) and the incidence of deformities was low (< 1%) in both groups. Marine survival was lower in the triploid group, largely as a consequence of higher losses sustained during a period of chronic stress, when triploid losses were 9% higher. Growth patterns were similar for the first 11 months in sea water. Although graded triploid salmon were heavier in January 1998 (AF3N 1.62 ± 0.033 kg, AF2N 1.46 ± 0.36 kg, P < 0.05), when the fish were harvested in May 1998 diploid salmon were significantly heavier than triploid salmon although there was no significant difference in weights after evisceration (AF3N 2.40 kg ± 0.04 AF2N 2.49 kg ±0.03). The increase in weight of the diploids between winter and harvest reflects the growth spurt that occurs in maturing fish in the spring. Overall yields of triploid salmon in salt water were lower as a result of inferior survival.     

Successful Hybridization of Acipenser Species Using Cryopreserved Sperm

Successful hybridization of sterlet (Acipenser ruthenus) × Siberian sturgeon (Acipenser baeri), sterlet × Russian sturgeon (Acipenser gueldenstaedti) and sterlet × European sturgeon (Acipenser sturio) was carried out for the first time using cryopreserved semen of sturgeon males and sterlet × sterlet crosses as controls. Sperm of all three species was diluted with a cryodiluent composed of 23.4 mM sucrose, 0.25 mM KCl, 30 mM Tris (pH 8.0) and 10% methanol. The samples were frozen in plastic straws in the vapor of liquid nitrogen at the height of 3 cm above the level of nitrogen for 3 min. Following thawing approximately 3000 sterlet eggs were fertilized with six straws of frozen-thawed sperm. The hatching rate with sterlet sperm was 30.6% while the hatching rate of A. ruthenus × A. baeri, A. ruthenus × A. gueldenstaedti and A. ruthenus × A. sturio hybrids was 50, 17.4 and 34%, respectively. Morphometric markers as well as random amplified polymorphic DNA (RAPD) assay was used to verify interspecific hybridization.     

Survival, growth and dressing traits of triploid hybrids between rainbow trout and three charr species

Triploid hybrids between female rainbow trout Oncorhynchus mykiss and male brook charr Salvelinus fontinalis, Arctic charr S. alpinus and lake charr S. namaycush, together with diploid and triploid rainbow trout controls from the same dams, were tested in freshwater farming up to their fourth year of life. All hybrids displayed lower survival rates than the controls, the weakest genotype being the Arctic charr hybrid. Mortalities were mostly observed at the embryonic and larval stages and at the adult stage as a consequence of male sexual maturation. Growth of all hybrids was hindered (compared with controls) during the first year, but only moderate differences remained after 3 years. Sexual maturation resulted in a weight inferiority of males in all genotypes. As to carcass traits, female hybrids displayed a slightly higher dressing percentage than female triploid rainbow trout, as a result of lower visceral losses. These results are discussed with reference to hybrid resistance to rhabdoviruses from the viewpoint of fish farming improvement.     

Comparative Growth and Survival of Reciprocal Hybrids Between Clarias gariepinus and Clarias anguillaris

Reciprocal, interspecific hybrids of Clarias gariepinus and Clarias anguillaris were produced via hypophysation and dry stripping. Two-week-old fry (0.06 ± 0.02 g) of the parental Clarias species and their interspecific hybrids were stocked in duplicated 1 m³ hapas suspended in outdoor earthen ponds (16 m²) for a rearing period of 56 days. Growth and survival of the hybrids was intermediate to the faster growing and better surviving C. gariepinus and the more slowly growing and more vulnerable C. anguillaris parental lines. The weight gain attained by C. gariepinus, C. anguillaris, C. gariepinus (♂)?×?C. anguillaris (♀), and C. anguillaris (♂)?×?C. gariepinus (♀) were 3.18 g, 2.99 g, 3.16 g and 3.05 g, respectively. Among hybrids, the growth of C. gariepinus (♂)?×?C. gariepinus (♀) was higher than the reciprocal C. anguillaris (♂)?×?C. gariepinus (♀). Heterosis for growth was a negligible 0.65% and for survival was negative (?5%). C. gariepinus appears to be the best species for aquaculture in terms of growth rate and survival.     

Induction of triploidy in large yellow crocker Pseudosciaena crocea (Richardson, 1846): effects of pressure shocks and growth performance in the first rearing year

The precociously sexual maturation in large yellow crocker Pseudosciaena crocea has become a serious problem. In an attempt to solve this problem, the production of sterile triploids could be an effective strategy. In this study, triploid P. crocea was obtained by subjecting fertilized eggs to pressure shock. Flow‐cytometry analysis was used to assess ploidy level. In terms of triploid rate and hatching rate, the optimal conditions of pressure shock for triploidy induction in P. crocea were 7500 psi for 3 min shock at 3 min after fertilization at 20 °C. With the application of these parameters, 100% triploid fish were produced. During the first rearing year, triploid P. crocea had a similar growth performance compared with its diploid counterpart before the age of 8 months and showed a significant advantage at the age of 10 and 12 months in body weight and body length (P<0.05). At the age of 12 months, the carcass weight of triploids was markedly higher than that of diploid control, and gonadal somatic index was significantly lower than that of their diploid control. During the first rearing year, survival in triploid group was 76.44%, inferior to its diploid control (83.21%).     

Maternal variation in juvenile survival and growth of triploid hybrids between female rainbow trout and male brown trout and brook charr

Related sib‐groups of rainbow trout × brown trout and rainbow trout × brook charr triploid hybrids and monospecific diploid and triploid rainbow trout controls were obtained from a common set of rainbow trout dams. On the basis of hybrid juvenile performances, 10 sib‐groups were selected and the corresponding diploid rainbow trouts were raised up to adult stage. Females from each group of rainbow trout were used to produce a second generation of hybrid progeny, the performances of which were analysed for grandmaternal variation and relation with first‐generation relatives. Results showed that hybrid traits (alevin yield and weight, survival and growth of fingerlings) were strongly influenced by maternal origin, and could be correlated to those of rainbow trout controls, but that maternal abilities had a low rate of inheritance. It was concluded that little improvement can be expected through selective breeding within parental populations.     

A comparative study of growth and feed utilization efficiency of sex-reversed diploid and triploid Nile tilapia, Oreochromis niloticus L.

Growth performance, survival and feed utilization of diploid (2n) and triploid (3n) sex‐reversed male and female Nile tilapia were evaluated at maintenance feeding (1% body weight (BW) day^?1), fixed feeding (3% BW day^?1) and apparent satiation feeding levels in a freshwater recirculation system comprised of thirty‐six 1‐m³ concrete tanks at the Asian Institute of Technology, Bangkok, Thailand. Triploid Nile tilapia (3n) was produced by subjecting fertilized diploid (2n) tilapia eggs to heat shock. After hatching, fish were sex‐reversed to all‐male and all‐female populations by oral administration of 17 α‐methyltestosterone (60 mg kg^?1 feed) and ethynylestradiol (100 mg kg^?1 feed) respectively. There was significantly higher growth with increased ration levels in both male and female groups. There were no significant differences in final BW, specific growth rate, survival rate, feed conversion ratio and protein efficiency ratio between diploid and triploid fish. Triploids had lower gonad weights than diploids, and this was particularly evident at the satiation feeding level. Triploid fish had a significantly higher apparent net protein utilization and percentage of gutted weight than diploids at all feeding levels. Higher protein utilization efficiency of triploids might be an advantage for commercial tilapia culture but further research is necessary to make such a conclusion.     

Daily patterns of ammonia nitrogen output of Siberian sturgeon Acipenser baeri (Brandt) of different body weights

The daily pattern of ammonia nitrogen outputs of five weight groups of Siberian sturgeon, Acipenser baeri (Brandt), was studied. Regardless of weight, there was a delay of 3 h between the start of feeding and the increase in ammonia output. For sturgeon reared at 17°C with continuous feeding, the total ammonia output levels (NH₄-N + NH₃-N) decreased from 530 mg kg^?1 per day for fish weighing 40 g, to 239 mg kg^?1 per day for fish weighing 1700 g. The study showed that continuous feeding is suitable in sturgeon farms in terms of ammonia loadings.     

Ploidy polymorphism and morphological variation among reciprocal hybrids of Pseudosciaena crocea (♀) × Miichthys miiuy (♂)

Diploid and triploid hybrids of large yellow croaker (Pseudosciaena crocea, Pc) and Miiuy croaker (Miichthys miiuy, Mm) were produced by interspecific hybridization. The result showed that Pc (♀) and Mm (♂) can be successfully crossed and hybrid offspring of normal hatch rate were produced. In contrast, a lower fertilize and hatch rate were produced for hybrids of Pc (♂) and Mm (♀), and the offspring died day 19 after hatching. To confirm the ploidy of hybrid offspring, we examined the karyotypes of offspring. The result of DNA relative content investigation and genetic relationship analysis demonstrated that the major hybrid offspring were hybrid diploid. In addition, a small number of hybrid were triploid and gynogenesis diploid. Morphological analysis based on the measurable traits data indicated the success of the interspecific crosses. The natural gynogenetic diploid large yellow croaker (PP), diploid hybrids (PM) and triploid hybrids (PPM) were closer to maternal parent (Pc) than paternal parent (Mm). This study provided genetic evidences of the chromosome, DNA content and morphological levels to support the successful establishment of the polyploidy hybrids of Pc (♀) and Mm (♂).     

Hybrid sturgeon ‘AL’ (Acipenser naccarii×Acipenser baeri) diets: the use of alternative plant protein sources

In this experimentation, corn gluten (CG) and pea meal (PM) were tested as potential protein sources in hybrid sturgeon ‘AL’. One hundred and twenty‐eight hybrid sturgeons ‘AL’ (Acipenser naccarii × Acipenser baeri) (initial body weight 364.8 ± 4.5 g) were utilized with four experimental diets and four replicates each; the experimental design was 4 × 4. Four isonitrogenous [crude protein=40% dry matter (DM)] and isoenergetic (gross energy=20 MJ kg^?1 DM) diets were formulated containing different levels of CG with or without PM and tested against a control diet that was fish meal (FM) based. Diets were as follows: CG55 contained 55% of corn gluten; diet CG55PM25 contained 55% CG and 25% PM; diet CG45PM25 contained 45% of corn meal and 25% of PM; and diet FM was control, based on FM. Fish fed with the PM diet showed lower values of feed conversion ratio (FCR) and specific growth rate (SGR) (FCR=4.53 ± 2.51 in the CG55PM25 diet; FCR=4.09 ± 1.45 in the CG45PM25 diet; SGR=0.20 ± 0.07 in the CG55PM25 diet; SGR=0.19 ± 0.11 in the CG55PM25 diet). The results of tissue proximate composition confirmed the results obtained from productive traits. This study indicates that CG meal but not PM could be utilized as a substitute of FM in hybrid sturgeon ‘AL’ nutrition.