William L. Shelton

A Temperature-Dependent Index of Mitotic Interval (τ0) for Chromosome Manipulation in Paddlefish and Shovelnose Sturgeon

Abstract A temperature-dependent measure of the mitotic interval (τ0) can help standardize chromosome manipulation in fish eggs. A tau unit (τ0) is the duration in minutes of one mitotic cycle during synchronous embryonic cleavage. It is measured over a range of temperatures, and the resulting relationship of τ0 to temperature can be used to anticipiate developmental events that are affected by temperature. Optimum induction of chromosome manipulation requires development of a specific treatment of egg shocking for each species, Timing of shock is a critical variable, but pretreatment incubation temperature affects the rate of development and thus the optimum absolute time for shocking. Mitotic intervals (τ0) are reliable indicators of developmental rates over normal temperatures for egg incubation, and thus can be used to estimate optimal times for chromosome manipulation, Mitotic intervals for paddlefish Polyodon spathula and shovelnose sturgeon Scaphirhynchus platorynchus were estimated by averaging th...

Broodstock development for monosex production of grass carp

Abstract Monosex female grass carp production has been accomplished with a breeding program that involves functionally sex-inverted males as broodstock. Between 1977 and 1982, 102 phenotypic males were produced by treating genetic females with methyltestosterone; 16 have sired progeny. Only females have been found among 1339 offspring sexed from 3 year classes.

Ionic composition and osmolality of paddlefish (Polyodon spathula, Acipenseriformes) seminal fluid

Changes in ionic composition as Na+,K+, Ca2+ and Mg2+, osmolality inseminal fluid, percentage of motile spermatozoaand velocity were investigated in response toCPP and different dosage of LHRHa. The lowestvelocity of sperm was observed after use CPPtreatment. The velocity of spermatozoa,significant main effect of the treatment(P < 0.0001) and the time of sperm collection(P < 0.0104) were evaluated. The osmolality ofseminal fluid was different betweenexperimental groups of LHRHa (48.0–62.7mOsmol.kg−1) and CPP (33.0–46.3mOsmol.kg−1) treatments. The osmolalitywas significantly higher on the first day andone-half, then declined on day three, rangingfrom 33.0 to 62.7 mOsmol.kg−1. Analysisof variance showed significant main effects ofthe treatment (P < 0.0001) and the time ofsperm collection (P < 0.0002) on the osmolalityof seminal fluid. The level of Na+ andK+ ion was different between experimentalgroups of LHRHa and CPP treatment. The highestconcentration of 11.11 mmol.l−1 wasobserved at Na+ ion. Then theconcentrations declined on the level 1.56, 0.52and 0.36 mmol.l−1 for K+, Ca2+and Mg2+ ions, respectively. There werehighly positive correlations between osmolalityof seminal fluid and dosage of LHRHa treatment(r = 0.84), velocity of spermatozoa andosmolality of seminal fluid (r = 0.57) andosmolality of seminal fluid and Na+concentration at seminal fluid (r = 0.70).Injection with LHRHa increased quality of spermas velocity of sperm, level of Na+,K+ and osmolality at seminal fluidcompared to CPP treatments.

Effects of ions on the motility of fresh and demembranate spermatozoa of common carp (Cyprinus carpio) and paddlefish (Polyodon spathula)

In the present study was summarize the factors activating and/or inhibiting the motility of intact and/or demembranated spermatozoa of common carp and paddlefish as teleostean and chondrostean models using methods of sperm demembranation. The movement of spermatozoa of cyprinids species, common carp, is influenced by osmotic pressure or high concentration of ions. The movements of spermatozoa of chrondrostean species, paddlefish, are under reciprocal control by the concentration of K+ and Ca2+.

Induced gynogenesis in black crappie.

Abstract This study reports the results of initial experiments on induced diploid gynogenesis in black crappie Pomoxis nigromaculatus. White bass Morone chrysops were an effective sperm donor for gynogenetic experiments with black crappies. White bass spermatozoa fertilized black crappie eggs, but hybrid larvae were not viable and died after hatching. In a series of experiments, we determined the ultraviolet (UV) light dose required to inactivate the sperm genome and the heat shock parameters needed to restore diploidy. Black crappie eggs were inseminated with white bass sperm irradiated with UV dosages ranging from 25 to 2,500 J/m2. A typical “Hertwig effect” in the yield of hatched larvae was observed with dosages greater than 100 J/m2. The genetic inactivation of paternal chromosomes was confirmed by flow cytometry measurement of DNA content in larval cells. Larvae resulting from sperm irradiated at a dose of 1,000 J/m2 were haploid. Time durations of mitotic interval (τ0) at different temperatures for...

OVARIAN DIFFERENTIATION IN COMMON CARP (CYPRINUS CARPIO) IN RELATION TO GROWTH RATE

Abstract Growth rate is influenced by density-dependent factors and in turn may affect the ontogeny of some systems. Population density gradients were established in common carp nursery ponds to develop differential growth rates. Ovarian development at the anatomical and cytological stages of differentiation were examined in relation to growth. Size at the end of 66 ± 2 and 155 ± 5-day periods was inversely related to population density, and the size/age relationship of differentiation of the ovary was affected. Mitotic gametogenesis was initiated sooner within populations with more rapid growth but generally at larger sizes than in populations with slower growth rates. Application of this phenomenon can be important to selection of appropriate size and/or age of common carp and to determine the required duration of androgen-induced sex reversal treatments.

Minimally Invasive Surgical Removal of Ovulated Eggs from Paddlefish

LUDEǨ STĚCH, OTOMAR LINHART*, WILLIAM L. SHELTON and STEVEN D. MIMS Fish Culture Hluboka nad Vltavou, Tyrsova 681, 373 41 Hluboka nad Vltavou, Czech Republic; Research Institute of Fish Culture and Hydrobiology, University of South Bohemia, 389 25 Vodňany, Czech Republic; Zoology Department, University of Oklahoma, Norman, Oklahoma 73019, USA; Aquaculture Research Center, Kentucky State University, Frankfort, Kentucky 40601, USA

Induced meiotic gynogenesis in shovelnose sturgeon

Gynogenesis was induced in three shovelnose sturgeon (Scaphirhynchus pIatorynchus) by heat shock after egg activation with UV-treated paddlefish (Polyodon spathula) milt. Ultraviolet dosage (J m−2) for the pooled milt samples was calculated using the following linear regression equation: Dosage = 2405.27 − 352.80X 19.78X2 (X = percent transmittance of milt). Activated eggs were incubated at 18 °C until shocking at 35 °C. Shock duration was applied at 0.05τ0 intervals from 0.15 to 0.40 τ0 (8.25 to 22.00 min post-fertilization; τ0 at 18 °C = 55 min). The highest yield of gynogenotes (16%) was observed at 0.25 τ0 for female 3, 10 % at 0.30 τ0 for female 2 and 12% at 0.35 τ0 for female 1. The percentage of viable gynogenotes responded quadratically to the tau index (τs/τ0) when shock treatments were applied. The higher yields of viable diploid sturgeon gynogenotes were achieved when eggs were heat shocked at embryological ages ranging from 0.25 to 0.35 τ0 (approximately 14 to 19 min post-activation at 18 °C). No viable hybrids were produced in the control fertilization of sturgeon eggs with intact paddlefish sperm which verified the gynogenetic origin of the offspring produced.

Chromosome set manipulations in the black carp

The mollusc-eating black carp (Mylopharyngodon piceus) has economic and health-care potential for biological control of nuisance aquatic molluscs. The present study investigates the production of gynogenetic-monosex and triploid-sterile populations of black carp. The goal was to provide a method which would eliminate unwanted biological and environmental impacts of introducing this exotic species into areas with nuisance mollusc infestation. Meiotic gynogenesis was induced by inseminating black carp eggs with UV-irradiated (800 Jm−2) sperm of common carp (Cyprinus carpio) or Japanese ornamental (koi) carp. Diploidy was restored through retention of the second polar body (2PB), by shocking activated eggs at 1–8 min post-fertilization (embryological age of 0.07–0.57τ0, a parameter defined by the cell cycle duration) at 1 min intervals, with heat-shocks (41.0±1.0 °C for 1 min) or pressure-shocks (7500–7600 psi for 1.5 min). Highest survival was found when embryos were heat-shocked 1.5–4.5 min post-fertilization (0.10–0.25τ0). The highest survival of free-swimming larvae from pressure-shocked eggs, was achieved at 7500 psi at 1–2 min post-fertilization (0.08–0.16τ0). Triploidy was induced by retention of 2PB following normal fertilization. Batches of 30 000 eggs were fertilized with intact sperm and pressure-shocked (6000–8500 psi for 1.5 min) 2 min post-fertilization (0.15–0.16τ0). The highest survival of triploid swim-up larvae was 5.1% in eggs shocked with 7500 psi. In random samples of individual larvae taken from each treatment, triploidy was analysed by cytofluorometry of the cellular DNA content. In DNA analysis performed in fingerlings (N≥15), 50% of the fish were triploids.

Fabrication of Silastic Implants for in Vivo Steroid Delivery in Fish

Abstract The use of hormones is a basic tool for managing reproduction in aquaculture. Delivery of a steroid over an extended period via implantation permits the manipulation of phenotypic sex for some fishes for which administration of the steroid in feed is not an option. An implant fabricated from Silastic materials and filled with 17α-methyltestosterone (MT) has been used to sex-reverse several species of fish. The fish must be large enough to accommodate the implant during the period of gonadal differentiation; the diffusion of 5 mg of MT will take place over a period of about 1 year.