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

Lead Research Organisation: University of Edinburgh
Department Name: The Roslin Institute

Abstract

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.

Technical Summary

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.

Planned Impact

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.

Publications

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Houston R (2017) Future directions in breeding for disease resistance in aquaculture species in Revista Brasileira de Zootecnia

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Palaiokostas C (2017) Genome-wide approaches to understanding and improving complex traits in aquaculture species. in CAB Reviews: Perspectives in Agriculture, Veterinary Science, Nutrition and Natural Resources

 
Description The key findings can be split into two areas: firstly the development of a novel genetic tool (a SNP chip) for oysters, and secondly the use of the SNP chip to discover new findings about resistance to herpes virus in oysters. To develop the SNP chip, we sequenced diverse populations of oysters and discovered millions of genetic variants in their genomes. We then created a combined species oyster SNP chip which contains around 50,000 filtered genetic variants (SNPs). This tool has been widely applied in various parts of the world, including Mexico, Australia and New Zealand. In our project, we also performed a large disease challenge experiment on juvenile oysters and sampled the resistant and susceptible oysters. We then genotyped them using the SNP chip, and showed that host resistance is heritable and controlled in part by a significant quantitative trait locus (QTL) on oyster chromosome 6. These results are being taken up by the industry, including oyster hatcheries in the UK and New Zealand, to breed for improved resistance to disease in oyster aquaculture.
Exploitation Route The oyster SNP database and SNP array will be useful for researchers in bivalve genetics, including those interested in evolutionary and conservation genetics, as well as the genetics of complex traits and animal breeding. Once genes underpinning resistance to Oyster Herpes Virus have been discovered, this will provide fundamental knowledge of genetic resistance mechanisms in oysters, and will be useful to select the most resistant oysters for breeding improved stock.
Sectors Agriculture, Food and Drink

URL https://www.ed.ac.uk/roslin/news-events/latest-news/oyster-farming-to-benefit-from-new-genetic-screeni
 
Description Pacific oysters are one of the most important aquaculture species globally, with production of over 600M tonnes per annum. However, production is seriously hampered by outbreaks of oyster herpes virus (OHV) which causes high levels of mortality in juvenile stock. Few prevention and treatment options are available, and selective breeding to improve innate resistance is therefore an important goal. In this project, our team developed a new genomic tool, called a SNP array, to enable high resolution studies of the genetic basis of resistance to this virus. The SNP array was created by high depth sequencing of many different populations of oyster and is one of the first of its kind for shellfish (Gutierrez et al. 2017, G3). The SNP array is being widely used by the shellfish genetics and aquaculture community, including scientists in New Zealand, Australia, Mexico, and throughout Europe. We have also used this array to map regions of the oyster genome that effect resistance to OHV, with a particularly interesting locus on oyster chromosome 6 that has a relatively large effect on resistance in commercially-sources oyster juveniles (Gutierrez et al. 2018, G3). The outputs of the research have been of interest to trade publications targeting the shellfish production industry (e.g. https://www.hatcheryinternational.com/research/genetic-advance-for-oyster-culture-aimed-at-improving-broods-1649 http://assg.org.uk/download/i/mark_dl/u/4006929760/4633637167/Grower%202018%20Jan.pdf ). We have also worked alongside UK aquaculture companies to enable application of genetic marker-enabled selection in oyster hatcheries, specifically working with Guernsey Sea Farms and SeaSalter. Finally, assisted by a BBSRC partnering award (BB/N022114/1), the Roslin team have teamed up with the Cawthron Institute in New Zealand to improve oyster breeding and genetics, specifically developing novel genomic methods for breeding resistance to OHV in their farmed oyster populations, further increasing the international impact of the outputs of this research project.
First Year Of Impact 2016
Sector Agriculture, Food and Drink
Impact Types Societal,Economic,Policy & public services

 
Description Advancing European Aquaculture by Genome Functional Annotation
Amount € 6,000,000 (EUR)
Funding ID 817923 
Organisation European Union 
Sector Public
Country European Union (EU)
Start 05/2019 
End 04/2022
 
Description BBSRC Other Countries Partnerships
Amount £22,500 (GBP)
Funding ID BB/N022114/1 
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 04/2016 
End 04/2019
 
Description Genomic and nutritional innovations for genetically superior farmed fish to improve efficiency in European aquaculture
Amount € 6,149,963 (EUR)
Funding ID 818367 
Organisation European Union 
Sector Public
Country European Union (EU)
Start 01/2019 
End 12/2022
 
Description Horizon 2020 Call H2020-SFS-2016-2
Amount € 7,000,000 (EUR)
Funding ID 727315 
Organisation European Union 
Sector Public
Country European Union (EU)
Start 05/2017 
End 04/2021
 
Description NERC Aquaculture Innovation Award
Amount £202,253 (GBP)
Funding ID NE/P010695/1 
Organisation Natural Environment Research Council 
Sector Public
Country United Kingdom
Start 01/2017 
End 12/2018
 
Description Research Grant - Ross Houston - Investigating genetic resistance to Bonamia in European flat oyster
Amount £165,026 (GBP)
Organisation Blue Marine Foundation 
Sector Charity/Non Profit
Country United Kingdom
Start 07/2018 
End 06/2020
 
Description Standard Grant - Ross Houston - AquaLeap: Innovation in Genetics and Breeding to Advance UK Aquaculture Production
Amount £403,285 (GBP)
Funding ID BB/S004343/1 
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 01/2019 
End 12/2021
 
Description Collaboration with Cawthron Institute in New Zealand 
Organisation Cawthron Institute
Country New Zealand 
Sector Charity/Non Profit 
PI Contribution This collaboration is built around a BBSRC partnering award in which I was the UK PI (BB/N022114/1 New Zealand - UK Paterning Award: Breeding for disease resistance in farmed oysters using genomic tools). The idea is to merge the genomic tools and skills that we have developed in the UK with the advanced shellfish selective breeding programmes that Cawthron run in New Zealand. Our research team has, for example, developed a high density SNP genotyping array for farmed oysters - the first of its kind for this species.
Collaborator Contribution Cawthron have world-leading selective breeding programmes for shellfish, including Pacific Oysters. One of the main target traits is to increase resistance to Oyster Herpes Virus (OHV) which can decimate stocks on oyster farms. They are providing our collaborative projects with access to extensive samples and data from their pedigreed oysters which have been challenged with OHV, in addition to their time to manage and run the project and provide intellectual input.
Impact Funded collaborative grant: NE/P010695/1 Application of genetic markers to improve resistance to herpes virus in commercial oyster populations This grant application builds on the BBSRC travel award and will provide funds for genotyping Cawthron's pedigreed oyster material with our high density SNP array developed under the BBSRC Aquaculture Initiative project BB/M026140/1 Investigation of Host Genetic Resistance to Oyster Herpes Virus using a High Density SNP Array. The downstream impact will be improved selective breeding for disease resistance in oysters, and transfer of much needed skills and expertise in advanced shellfish breeding from NZ to the UK. These collaborations also involve the Centre for Environment, Fisheries and Aquaculture Science (Cefas). The collaboration brings together expertise in genomics, sequencing, selective breeding, shellfish biology and virology.
Start Year 2016
 
Description Strategic research partnership with WorldFish 
Organisation Worldfish
Country Malaysia 
Sector Charity/Non Profit 
PI Contribution A strategic research partnership has been established between Roslin and WorldFish. This partnership is initially focussed on use of advanced genetic and genomic technology to improve selective breeding of tilapia - one of the world's most important foodfish. Our contribution has included development of a research programme that builds on research performed in Atlantic salmon to inform strategies and techniques to implement genomic selection in WorldFish tilapia breeding programme.
Collaborator Contribution WorldFish run a family based selective breeding programme for tilapia and will provide data and samples from this programme to inform the research.
Impact This is a multidisciplinary partnership involving researchers involved in tilapia health, genetics, molecular biology and bioinformatics.
Start Year 2017
 
Description Organised a British Council Researcher Links conference in Mexico 
Form Of Engagement Activity A formal working group, expert panel or dialogue
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Co-organised a Researcher Links conference about Genomics in Aquaculture in Merida, Mexico in January 2017. This 5 day event featured around 40 scientists, with a mix of young researchers and mentors from both the UK and Mexico. This has already led to several collaborative links between UK and Mexico in this field.
Year(s) Of Engagement Activity 2017
URL https://sites.google.com/site/genomicsinaquaculturemxuk/home