Development of Novel and Emerging Technologies for Cryogenic or Long-term Preservation and Revival of Drosophila and Zebrafish Genetic Stocks (R43/R44 Clinical Trial Not Allowed).

Animal models are essential for research, contributing to the understanding of the pathobiology of human diseases and facilitating the development of diagnostic approaches and therapeutic interventions. A large number of animal genetic stocks, including models of human diseases, have been generated at an unprecedented fast pace due to rapidly evolving gene editing technologies. However, this rapid increase in genetic stocks is creating challenges in sustainability of the resources, i.e., how to maintain and preserve these critical resources in reliable, cost-effective and long-term ways. The National Institutes of Health (NIH) has invested a substantial amount of funding to generate and maintain these resources in dedicated stock centers that serve the biomedical research community. The fruit fly (Drosophila melanogaster) and the zebrafish (Danio rerio) are two of the leading non-mammalian model organisms used for the understanding of basic biology of animal systems and modeling human diseases; however, cryogenic approaches in these species have not been fully established and, in many cases, have proven to be unsuccessful. Drosophila and zebrafish have become increasingly important as biological resources because of their small size, relatively short generation time, easy manipulation of embryos and optical transparency as well as genetic tractability. Both are important for studies in diverse scientific areas, including genetics, pharmacology, toxicology, neurobiology, behavior and developmental biology. Currently there are over 60,000 Drosophila stocks at the NIH-supported Bloomington Drosophila Stock Center and nearly 11,000 fish-lines (live and preserved) at the Zebrafish International Resource Center, respectively, but these numbers are increasing at a rapid pace. There is an urgent need to achieve simple, reliable, reproducible and cost-effective methods for cryogenic or long-term preservation and revival of Drosophila and zebrafish stocks, where preservation options are essentially absent (fruit fly) or limited (zebrafish), as compared to the cryopreservation methods developed and widely used for several other animal models, such as worms, mice and rats; and to ensure efficient, and reproducible maintenance and revival of their genetic stocks.

ORIP convened two workshops to offer a forum for discussion of major scientific developments and road blocks that prevent preservation of any stage of fruit fly or beyond sperm for zebrafish (see Cryopreservation of Drosophila Strains and Cryopreservation of Aquatic Biomedical Models Workshop Final Reports). Taking into consideration the recommendations of participants at both workshops, ORIP encourages SBIR applications aimed at developing and improving technologies for cryogenic or other long-term preservation and revival of Drosophila and zebrafish stocks, including, but not limited to, the following:

• Development of new cryopreservation/preservation germplasm procedures and tools to achieve efficient, reliable, reproducible and cost-effective methods for genetic banking, including vitrification and pipelines specially for embryos, eggs, and oocytes of Drosophila or zebrafish;
• Specialized reagents to minimize cellular damage to a wide variety of stocks during collection, processing, storage and revival. Examples include developing reagents for vitrification or drying of germplasm (Drosophila/zebrafish) or larvae (Drosophila);
• Novel methods for cryopreservation and dehydration of implantable gonad tissue (ovaries or testes), germplasm (Drosophila/zebrafish) or larvae (Drosophila);
• Tools and processes to standardize cryogenic, long-term or medium-term preservation protocols that will increase the reliability, reproducibility, simplicity, throughput, and efficiency of stock preservation;
• Biosecurity methods to diagnose and eliminate pathogens and opportunistic pathogenic microorganisms that may contaminate embryos or other types of germplasm (Drosophila/zebrafish) or larvae (Drosophila);
• High throughput technologies and microfluidics devices that can speed up the long-term preservation and revival of embryos, eggs, sperm, oocytes, stem cells, and somatic cells;
• High throughput systems to speed up phenotypic and toxicological analysis using zebrafish embryos, particularly technologies to automate dispensing and orienting of embryos in multi-well plates;
• Identification and detection of biological markers to evaluate the status of preservation and revival of embryos, eggs, sperm, oocytes, stem cells, somatic cells (Drosophila/zebrafish) or larvae (Drosophila).

Applicants are strongly encouraged to discuss their projects with the Scientific/Research contacts before submitting an application.

See Section VIII. Other Information for award authorities and regulations.