Name: Video: Computer Assisted Coral Sperm Analysis and Cryopreservation for Biobanking
Uploaded: 2024-06-07T13:30:00
Duration: 4 min 6 s
Description: 74 Views. Computer Assisted Coral Sperm Analysis and Cryopreservation for Biobanking

computer assisted coral sperm analysis and cryopreservation for biobanking

Optimizing Coral Sperm Cryopreservation from Threatened Coral Species

Coral reefs globally are experiencing a loss of coral species, populations, and genetic diversity due to ocean warming and acidification caused by climate change, diminishing the viability of these critical habitats and impacting the species that they support1,2. The most effective approach to secure genetics long-term is cryopreservation and biobanking, which permits the frozen storage of living samples at cryogenic temperatures in liquid nitrogen indefinitely3. The development in 2012 of methods for the cryopreservation of coral sperm4 enabled the biobanking of genetics from these species for the first time and led to the development of the first biorepository for coral genetics in 20125. Since then, this cryopreservation protocol has been further refined6 and used to secure the genetics of over 50 species of coral globally, with 30 of those species coming from the Great Barrier Reef in Australia7. Cryopreserved coral sperm can generate healthy corals that develop normally and have been used to facilitate assisted gene flow experiments in Australia8 and the Caribbean9. While technologies are currently under development to enable the cryopreservation of complex tissue types such as larvae10 and adult coral tissues11,12, cryopreservation of coral sperm is currently the most established tool available for routine biobanking of coral genetics.
As the impacts on coral populations have increased, several countries have initiated large-scale programs to support reef restoration and adaptation (e.g., the Reef Restoration and Adaptation Program [RRAP] in Australia13) or to secure remaining imperiled coral populations (e.g., Florida Coral Rescue in the USA14). In the context of these programs, cryopreservation can be thought of as an enabling technology, supporting research and large-scale coral production in addition to securing existing genetic diversity and preventing extinctions. Sperm cryopreservation can enable greater control over reproduction between populations that are physically or temporally separated and can permit genetic management of broodstock to select for desirable traits such as thermal tolerance or disease resistance8. To date, coral sperm biobanking has been undertaken on a relatively small scale in support of biodiversity management7, so some level of upscaling will be required if such biobanking is to meet its potential within these larger reef restoration programs. As with all reef restoration efforts based on coral reproduction, the main impediment to increasing biobanking efforts is the limited period during which coral gametes are available, since spawning in most reef-building species coincides with the full moon in late spring and early summer15, meaning that gametes are only available for cryopreservation and biobanking on a few nights each year. Moreover, in regions where mass spawning occurs, e.g., the Great Barrier Reef, there are typically multiple species spawning simultaneously or within a few hours of each other on the same night15. Enhancements to the sperm cryopreservation pathway are therefore required to increase the scale and efficiency of processing to maximize biobanking capacity during these brief annual spawning windows while ensuring the integrity of samples and associated metadata.
Cryopreservation and biobanking of coral sperm involve several key steps, from the collection of gametes during spawning to the accession of samples into the biorepository and database (Figure 1). The process begins with the separation of sperm from eggs (in hermaphroditic species) or the collection of sperm from the water column (in gonochoric species), followed by the assessment of sperm motility and concentration. Sperm are then combined with cryoprotectant (10% v/v final concentration, dimethyl sulfoxide, DMSO) in filtered seawater (FSW) and cooled at &#8722;20 &#176;C/min in a custom-designed cooling device4. Early iterations of this process relied on visual assessment of sperm motility and concentration via phase-contrast microscopy and hemocytometer counts, printing, and labeling tubes as samples were processed for cryopreservation, hand-pipetting sperm samples into cryovials, and cooling on a specially designed floating rack16. The application of computer-assisted sperm analysis (CASA)17 and the development of a 3D-printed cooling device6 have improved the efficiency and reliability of coral sperm cryopreservation, but the sperm sample processing pathway has remained largely the same since its inception. While this approach is suitable for processing samples from a moderate number of colonies and species on a spawning night, for large volumes and numbers of colonies (e.g., individual samples from &#62;10 colonies at a time), there are bottlenecks in the cryopreservation pathway that impede processing of the samples in a timely manner (i.e., within 2 h of sperm release) without degradation of sample fertility potential. Although large-scale fluid handling systems for cryovials are available (e.g., Thomas et al.18), they are typically designed for large batch processing (i.e., multiple racks of cryovials) and are not suitable for transport and use in the field, so they are not cost-effective for this application. Therefore, the present study aimed to improve the efficiency of coral sperm cryopreservation by introducing portable, inexpensive automation equipment and methods at key steps in the processing pathway to maximize the number of samples that could be effectively cryopreserved on a spawning night.

Author Spotlight: Developing Efficient Cryopreservation and Biobanking Technologies for Global Reef Restoration

A Semi-Automated Workflow for the Cryopreservation of Coral Sperm to Support Biobanking and Aquaculture

Computer Assisted Coral Sperm Analysis and Cryopreservation for Biobanking

computer-assisted-coral-sperm-analysis-cryopreservation-for

Research

JoVE Journal

Biology

74 Views. Computer Assisted Coral Sperm Analysis and Cryopreservation for Biobanking

Video: Computer Assisted Coral Sperm Analysis and Cryopreservation for Biobanking

Coral reefs are facing a crisis as the frequency of bleaching events caused by ocean warming increases, resulting in the death of corals on reefs around the world. The subsequent loss of genetic diversity and biodiversity can diminish the ability of coral to adapt to the changing climate, so efforts to preserve existing diversity are essential to maximize the resources available for reef restoration now and in the future. The most effective approach to secure genetics long-term is cryopreservation and biobanking, which permits the frozen storage of living samples at cryogenic temperatures in liquid nitrogen indefinitely. Cryopreservation of coral sperm has been possible since 2012, but the seasonal nature of coral reproduction means that biobanking activities are restricted to just a few nights per year when spawning occurs. Improving the efficiency of coral sperm processing and cryopreservation workflows is therefore essential to maximizing these limited biobanking opportunities. To this end, we set out to optimize cryopreservation processing pathways for coral sperm by building on existing technologies and creating a semi-automated approach to streamline the assessment, handling, and cryopreservation of coral sperm. The process, which combines computer-assisted sperm analysis, barcoded cryovials, and a series of linked auto-datasheets for simultaneous editing by multiple users, improves the efficiency of both sample processing and metadata management in the field. Through integration with cross-cutting research programs such as the Reef Restoration and Adaptation Program in Australia, cryopreservation can play a crucial role in large-scale reef restoration programs by facilitating the genetic management of aquaculture populations, supporting research to enhance thermal tolerance, and preventing the extinction of coral species. The described procedures will be utilized for coral cryopreservation and biobanking practitioners on reefs worldwide and will provide a model for the transition of cryopreservation technologies from research laboratories to large-scale applications.

This protocol describes a semi-automated pathway to improve the efficiency and capacity of processing and cryopreservation of sperm from threatened coral species, aiming to secure genetic diversity and support reef restoration efforts.

Coral reefs are facing a crisis as the frequency of bleaching events caused by ocean warming increases, resulting in the death of corals on reefs around ...