211 research outputs found
Strategies for commercialization of cryopreserved fish semen
No full textInitial success in sperm cryopreservation occurred at about the same time for aquatic species and livestock. However, in the 50 plus years since then cryopreserved sperm of livestock has grown into a billion-dollar global industry, while cryopreserved sperm of aquatic species remains a research activity with little commercial application despite work in more than 90 species and more than 200 published reports. Most research work has focused on large-bodied culture and sport fishes, such as salmon, trout, carp, and catfish, and mollusks such as commercially important oyster and abalone species. However, only a few studies have addressed sperm cryopreservation in small fishes such as zebrafish, or in endangered species. Overall, this work has yielded techniques that are being applied with varied levels of success around the world. Barriers to expanded application include a diverse and widely distributed literature base, technical problems, small sperm volumes, variable results, a general lack of access to the technology, and most importantly, a lack of standardization in practices and reporting. The benefits of cryopreservation include at least five levels of improvements for existing industries and for creation of new industries. First, cryopreservation can be used to improve existing hatchery operations by providing sperm on demand and simplifying the timing of induced spawning. Second, frozen sperm can enhance efficient use of facilities and create new opportunities in the hatchery by eliminating the need to maintain live males, potentially freeing resources for use with females and larvae. Third, valuable genetic lineages such as endangered species, research models or improved farmed strains can be protected by storage of frozen sperm. Fourth, cryopreservation opens the door for rapid genetic improvement. Frozen sperm can be used in breeding programs to create improved lines and shape the genetic resources available for aquaculture. Finally, cryopreserved sperm of aquatic species will at some point become an entirely new industry itself. A successful industry will require integrated practices for sample collection, refrigerated storage, freezing, thawing, rules for use and disposal, transfer agreements, and database development. Indeed the development of this new industry is constrained by factors including the technical requirements for scaling-up to commercial operations during the transition from research, and the absence of uniform quality control practices, industry standards, and appropriate biosecurity safeguards
Willingness-to-Pay for Genetic Attributes in Aquaculture Industries
No full textThe genetic make-up of fish stocks is an important factor in aquaculture production. Choice-based conjoint analysis is used to determine importance of genetic improvements to grow-out producers and an estimated willingness-to-pay for selected attributes. Results from a national survey of aquaculture producers, reveal growth rate as the most important attribute.Resource /Energy Economics and Policy,
Standardized Assessment of Thin‐film Vitrification for Aquatic Species
No full textUltrarapid cooling under the appropriate conditions will produce vitrification, a glass-like state used to cryopreserve small sample volumes, but there are a number of major technical drawbacks impeding the application of vitrification to germplasm of aquatic species. These include a lack of suitable devices, and poor reproducibility and comparability among studies due to a lack of standardization. We used 3-D printing to produce a viewing pedestal coupled with a classification system to rapidly assess frozen film quality of vitrification loops. Classification time declined with practice from 2.1 ± 0.3 sec (mean ± SD) to 1.5 ± 0.2 sec (after 200 assessments), and assessments were consistently made in \u3c 2.5 sec. Classifications should be reported with representative images allowing harmonization for quality control. This approach permits rapid classification and can be applied for the development of methods including the evaluation of vitrification solution components, concentrations of solutions and target cells, and configurations and volumes of new devices. Future studies should address the custom fabrication of 3-D printed vitrification devices for use with aquatic species and other applications
Environmental salinity-induced shifts in sperm motility activation in Fundulus grandis
No full textMotility activation of fish sperm typically responds to levels of specific ions or osmotic pressure differences between the surrounding water and body tissues. In general, the sperm of marine fishes are activated by an increase in osmotic pressure (hypertonic salinity), and that of freshwater species by a decrease (hypotonic salinity). These stenohaline species exist in relatively stable environments, however, estuarine fishes are exposed to rapidly changing and broad salinity ranges, often resulting in external osmotic pressures that include those of the body (isotonic). To assess the ability of Fundulus grandis sperm to adapt to changes in salinity, adult males were acclimated to salinities of 0, 5, 10, 20, 35, or 50ppt and held for 30d. The testes were dissected from the fish and sperm were activated with deionized water, various osmolalities (100-1000mOsmol/kg) of Hanks\u27 balanced salt solution (HBSS), calcium-free HBSS (Ca 2+-Free HBSS), and sodium chloride solution (NaCl). The deionized water did not activate sperm motility regardless of the acclimated salinity. Compared to HBSS, Ca 2+-Free HBSS and NaCl activated sperm motility with a significantly lower percentage at the same osmolalities. The osmolality eliciting the highest motility activation was significantly different (P\u3c0.01) among acclimated groups and shifted from 300mOsmol/kg (ranging from 200 to 500) for sperm collected from 5ppt, 500mOsmol/kg (ranging from 200 to 800) for sperm collected from 10ppt, 600mOsmol/kg (ranging from 400 to 700) for sperm collected from 20ppt, 800mOsmol/kg (ranging from 200 to 900) for sperm collected from 35ppt, and 900mOsmol/kg (ranging from 600 to 1000) for sperm collected from 50ppt. Motility peaked after 30s exposure to HBSS, and decreased over 10min. Motility exhibited a similar initial pattern when exposed to Ca 2+-Free HBSS, however, the sperm gained motility at lower osmolalities over 10min, exhibiting multiple peaks. These results indicate that environmental salinity can significantly influence sperm behavior in adult males of F. grandis with substantial changes after only 30d of acclimation. As such, this should be considered as a major unrecognized variable in sperm research in this species and can be considered for use in optimizing protocols addressing in-vitro fertilization and cryopreservation. Whether this phenomenon is unique to Fundulus or is a characteristic of euryhaline fishes remains unresolved. © 2011 Elsevier B.V
Strategies for commercialization of cryopreserved fish semen
No full textInitial success in sperm cryopreservation occurred at about the same time for aquatic species and livestock. However, in the 50 plus years since then cryopreserved sperm of livestock has grown into a billion-dollar global industry, while cryopreserved sperm of aquatic species remains a research activity with little commercial application despite work in more than 90 species and more than 200 published reports. Most research work has focused on large-bodied culture and sport fishes, such as salmon, trout, carp, and catfish, and mollusks such as commercially important oyster and abalone species. However, only a few studies have addressed sperm cryopreservation in small fishes such as zebrafish, or in endangered species. Overall, this work has yielded techniques that are being applied with varied levels of success around the world. Barriers to expanded application include a diverse and widely distributed literature base, technical problems, small sperm volumes, variable results, a general lack of access to the technology, and most importantly, a lack of standardization in practices and reporting. The benefits of cryopreservation include at least five levels of improvements for existing industries and for creation of new industries. First, cryopreservation can be used to improve existing hatchery operations by providing sperm on demand and simplifying the timing of induced spawning. Second, frozen sperm can enhance efficient use of facilities and create new opportunities in the hatchery by eliminating the need to maintain live males, potentially freeing resources for use with females and larvae. Third, valuable genetic lineages such as endangered species, research models or improved farmed strains can be protected by storage of frozen sperm. Fourth, cryopreservation opens the door for rapid genetic improvement. Frozen sperm can be used in breeding programs to create improved lines and shape the genetic resources available for aquaculture. Finally, cryopreserved sperm of aquatic species will at some point become an entirely new industry itself. A successful industry will require integrated practices for sample collection, refrigerated storage, freezing, thawing, rules for use and disposal, transfer agreements, and database development. Indeed the development of this new industry is constrained by factors including the technical requirements for scaling-up to commercial operations during the transition from research, and the absence of uniform quality control practices, industry standards, and appropriate biosecurity safeguards
Identification of sex in chickens by flow cytometry
No full textIn chickens, the difference in DNA content between the Z and W sex chromosomes is measurable as a 2% greater amount of DNA in cells of males (ZZ) vs. females (ZW). High-resolution measurement of cellular DNA content (genome size) is possible by use of flow cytometry. In this technique, nuclei are stained with a nucleic-acid specific fluorochrome, and are passed single-file through a high-intensity light source such as a laser beam. The fluorescence emitted from each nucleus is directly proportional to the amount of DNA present, and can be used to quantify genome size on a per-cell basis. This technique is rapid (two or three samples per minute after a 10-minute staining period), accurate, and straightforward. Commercially available clinical and research instruments are generally expensive (US$80,000 to 400,000), but it may be possible to couple the principles of flow cytometry with existing egg handling equipment to enable rapid throughput for identification of sex in chicks before hatching
Construction of an inexpensive low-temperature incubator
No full textA simple and inexpensive procedure is described for the modification of standard compact refrigerators for use as incubators at temperatures between 0°C and room temperature. © 1993 by the American Fisheries Society
Three-Dimensional Printing of Vitrification Loop Prototypes for Aquatic Species
No full textVitrification is a method of cryopreservation that freezes samples rapidly, while forming an amorphous solid ( glass ), typically in small (μL) volumes. The goal of this project was to create, by three-dimensional (3D) printing, open vitrification devices based on an elliptical loop that could be efficiently used and stored. Vitrification efforts can benefit from the application of 3D printing, and to begin integration of this technology, we addressed four main variables: thermoplastic filament type, loop length, loop height, and method of loading. Our objectives were to: (1) design vitrification loops with varied dimensions; (2) print prototype loops for testing; (3) evaluate loading methods for the devices; and (4) classify vitrification responses to multiple device configurations. The various configurations were designed digitally using 3D CAD (Computer Aided Design) software, and prototype devices were produced with MakerBot ® 3D printers. The thermoplastic filaments used to produce devices were acrylonitrile butadiene styrene (ABS) and polylactic acid (PLA). Vitrification devices were characterized by the film volumes formed with different methods of loading (pipetting or submersion). Frozen films were classified to determine vitrification quality: zero (opaque, or abundant crystalline ice formation); one (translucent, or partial vitrification), or two (transparent, or substantial vitrification, glass). A published vitrification solution was used to conduct experiments. Loading by pipetting formed frozen films more reliably than by submersion, but submersion yielded fewer filling problems and was more rapid. The loop designs that yielded the highest levels of vitrification enabled rapid transfer of heat, and most often were characterized as being longer and consisting of fewer layers (height). 3D printing can assist standardization of vitrification methods and research, yet can also provide the ability to quickly design and fabricate custom devices when needed
Amine reactive dyes: An alternative to estimate membrane integrity in fish sperm cells
No full textFluorescent dyes that binds irreversibly to cellular amines, come in several available emission spectra, and do not poses health concerns were used to evaluate membrane integrity in fish sperm cells. The objectives of the present study were to determine: (1) a working dye concentration for fish sperm samples, and (2) if the traditional propidium iodide/SYBR-14 staining combination was comparable with the amine reactive dye (ARD) methods at identifying cell populations with intact and compromised membranes after sperm activation, refrigerated storage, and exposure to cryoprotectant and surfactant. Zebrafish (Danio rerio) sperm were obtained by stripping, and pooled samples (in triplicate) were used in all tests. Six dilutions of the amine dye (ranging from 0.625 to 0.02 μl/ml) were evaluated, and compared with the traditional staining protocol. A concentration of 0.5 μl/ml ARD was selected to be used in subsequent assays.Sperm suspensions were activated with deionized water to simulate urine contamination. After 10 s, osmolality was increased to stop activation, and the procedure was repeated in 10-s intervals until the sperm remained activated for 120 consecutive sec; membrane integrity was analyzed at each time interval. For the storage assay, sperm suspensions were prepared in Hanks\u27 balanced salt solution at 302 mOsm/kg osmolality (HBSS302), HBSS354 and HBSS402, and evaluated every 2 h for 8 h, and every 24 h for 72 h. Cryoprotectant toxicity was tested by diluting sperm suspensions in HBSS340 with methanol at 5, 10 and 15% final concentrations. Surfactant toxicity was tested by diluting sperm suspensions in HBSS354 with Triton X-100 at 0.2, 0.15 and 0.1 mM final concentrations. In each toxicity assay, membrane integrity was tested every 20 min for 80 min. The number of membrane-intact cells significantly decreased across time in all treatments (P \u3c 0.05). Significant differences between staining protocols were observed after activation and after exposure to methanol at 10 and 15%, and to Triton X-100 (P \u3c 0.05). The average difference, however, was minor (between 1 and 6% in average) in relation to the typical values used for decision making based on this assay. Results showed that this method has the potential to contribute greatly to the standardization of cryopreservation in aquatic species. Statement of relevance: QA-QC for repository development in aquatic species
Concepts, History, Principles, and Application of Germplasm Cryopreservation Technology
No full textGermplasm are living genetic resources that can serve as bearers of heredity, and include germ cells and their precursors, plant seeds and pollen, animal sperm, oocytes, embryos, and larvae. Cryopreservation refers to the preservation of biological materials at extremely low temperatures, typically using solid carbon dioxide at -80°C or liquid nitrogen at -196°C for freezing, and cryogenic storage in perpetuity. Germplasm cryopreservation is an important technology applied for medical treatment, maintenance of biological diversity, preservation of valuable genetic resources, assistance of breeding programs, and conservation of imperiled species. This extension publication is intended to introduce the basic concepts, history, principles, and applications of germplasm cryopreservation technology
- …
