Investigation of Host Genetic Resistance to Oyster Herpes Virus using a High Density SNP Array

Pacific oyster (Crassostrea gigas) is one of the most important aquaculture species in the world, with an annual production of >0.6 billion tonnes. Due to its high growth rate and tolerance of a wide range of environments, C. gigas is farmed in most regions of the world, including the UK. However, Oyster herpes virus (OsHV-1) - a double-stranded DNA virus - has become the primary concern for C. gigas farming, often causing huge losses to farmed stocks in what is known as 'summer mortality syndrome'. The negative impact of the viral outbreaks on oyster production has been worsened by the emergence of a more pathogenic form; namely OsHV-1 micro variant. While restricting the movement of infected stocks and other biosecurity measures have had limited success in controlling outbreaks, recent studies suggest that there is genetic variation in the resistance of the oyster to the virus.

While many genomic tools and resources exist for C. gigas, there is not yet a high density genotyping platform suitable for detailed studies into the genetics of complex traits, such as disease resistance. Single nucleotide polymorphism (SNP) arrays are widely used in farmed terrestrial livestock and fish to research the genetic control of economically important traits. The genetic marker information can be applied to predict which animals are suitable for breeding programs to improve these traits. Additionally, these SNP arrays allow management of the diversity of population, and evolutionary genetic studies in species of interest.

Therefore, the two major aims of the proposed project are; firstly, to develop and test the first high-density SNP genotyping array for oysters and; secondly, to quantify the genetic variation in the resistance of oysters to herpes virus and determine whether this variation includes loci of major effect.
The first aim will be achieved by sequencing the genomes of a wide panel of oysters and mining the sequence data for SNPs. The SNP data will be used to develop a microarray containing approximately 50,000 individual SNP assays dispersed across the oyster genome. The second aim will be achieved by crossing 30 individual parent oysters taken from an oyster producer in Guernsey and challenging the offspring with a calibrated dose of the micro form of herpes virus. Samples of the oysters will then be taken and used for estimating the level of infection for each oyster and also for extracting DNA. This DNA will be genotyped using the new SNP array and the genome-wide marker data will be analysed to determine how much variation in viral resistance is genetic, and to map any major resistance loci within the oyster genome.

From the results of this project, it will be possible to use genetic markers to predict whether an individual oyster is likely to be resistant or susceptible to herpes virus. This information can be incorporated into selective breeding programs to help tackle this major disease threat. By mapping the loci affecting resistance to the genome of the oyster, important resistance genes may be identified and studied in future projects. Additionally, the SNP array tool developed will have wide applications for oyster genetics, including managing genetic diversity or differentiating between oyster species. As a final output from the project, we will encourage collaborative research in conservation or ecological genetics comparing different wild oyster populations; we anticipate having excess arrays available from this project and we will offer these at no cost to interested collaborators.

Pacific oyster (C. gigas) is one of the most extensively farmed aquaculture species in the world. Selective breeding programmes are at a formative stage, and have been established to improve disease resistance. The virulent 'micro' variant of Oyster Herpes Virus (OsHV-1) is the primary disease concern for oyster production, causing mass mortality in juvenile oysters. Estimates of the heritability of host resistance to the virus are moderate to high, which raises the possibility of identifying and breeding from resistant stock. To assist with this goal, a high density SNP array would enable thorough investigation of the genetic architecture of host resistance in disease-challenged families. Additionally, such an array would facilitate population genetic studies and monitoring of genetic diversity in wild oysters.

The two major aims of the proposed project are (i) to develop and test the first high-density SNP genotyping array for oysters and (ii) to quantify the amount and architecture of genetic variation in the resistance of oysters to OhHV-1. To achieve this, high coverage whole genome sequencing of pools of oysters from diverse populations will be performed. Sequence data will be aligned to the C. gigas genome and abundant SNPs discovered. These SNPs will be systematically filtered to create an Affymetrix Axiom array containing ~50 K markers. Experimental challenges of oyster families with OsHV-1 micro variant will be performed, with collection of survival data and samples, and measurements of viral load. These samples will be genotyped using the newly created SNP array and genome-wide marker data will be analysed to determine how much variation in viral resistance is genetic, and to map any major resistance loci within the oyster genome. The results will enable selective breeding for improved resistance, contributing to an effective control strategy for the disease, and provide a novel tool for future genetics studies in oyster species.

The primary goal of this project is to tackle Oyster Herpes Virus (OsHV-1), the primary threat to the oyster aquaculture industry worldwide, via improvements in host genetic resistance. As part of the proposed research program, we will also develop the first high density SNP genotyping array for oyster species, which provides a novel tool which can be utilised by the academic and commercial community for high-resolution genetics research in oysters. Our links with the UK oyster aquaculture industry mean that, if successful, the outcomes of this project are likely to be immediately and enthusiastically translated into practice for immediate positive economic impact. There will also be downstream beneficial impact for the scientific community via the tools and knowledge developed within the project, particularly the new SNP array. Finally, the general public and policy makers will benefit from improved efficiency of oyster production and downstream applications of the SNP array to monitor and preserve biodiversity, e.g. management of native stocks of UK flat oyster.

Industry: OsHV-1, in particular the more virulent 'micro' variant, is the single largest disease threat to the oyster aquaculture industry in the UK and worldwide due to massive mortality outbreaks. The virus is closely linked with water temperature, suggesting that warmer waters associated with climate change will increase the threat posed by the disease. Therefore, routes to tackle OsHV-1 are an industry priority. This project will result in knowledge of the genetic basis of host resistance and, critically, genetic marker predictors of resistance. The team will work with one of the UK's largest and technically-advanced producers, Guernsey Sea Farms, to ensure effective translation of results to a commercial setting. This is likely to lead to a multi-marker genetic test that can be used to inform breeding decisions by predicting whether oysters are resistant or susceptible from a DNA sample. The novel SNP array developed in the project may also have more general applications for parentage assignment and marker-based selection as oyster breeding programmes become more advanced.

Scientific Community: The project outcomes will be of considerable interest and value for researchers in related fields. The first and most obvious impact will be fundamental knowledge of the host response to herpes virus in oysters. Since disease challenge experiments in aquaculture species tend to be undertaken on a scale that is not practical / possible in other species, the results may have comparative value for the host response to herpes viruses generally. The SNP array will contain information from both C. gigas and O. edulis species and the data generated will vastly improve genomic resources for communities studying either of these species. Through links with scientific communities interested in population genetics of oyster species (see CACHE letter of support) we anticipate wide uptake of the SNP array tool for genetic studies in wild populations. Further, the project will provide substantial training for the PDRA as well as project management development opportunities for the team as a whole.

General Public and Policy Makers: The improvement of the sustainability of oyster production will lead to a more reliable source of this high quality product, potentially resulting in health benefits to society. The SNP array tool will enable effective monitoring and ultimately conservation of the European flat oyster O. edulis in its native environment and has the potential to contribute to the control of 'invasive' C gigas populations. Government policy makers are likely to benefit from the research through its contribution to a sustainable aquaculture industry. The research will be communicated to the public via interaction with the media, presentations, publications, exhibitions and schools activities - supported by a policy of clear and open communication and public engagement.