Algal vaccines for Aquaculture

Lead Research Organisation: University College London
Department Name: Biochemical Engineering

Abstract

The global salmon industry has grown rapidly over the last four decades, and the majority of salmon purchased by consumers is farmed. Therefore, given the value of the market, protection of farmed fish against diseases is of enormous importance. The salmonid alphavirus (SAV), is reported to be one of the most prominent and detrimental diseases in Irish, Scottish and Norwegian aquaculture industries. Infection of farmed fish causes substantial losses in production and revenue. At present, the most effective method for protecting salmon stocks is via injection of fish. However, his is hugely labour intensive and can induce other side effects.
Administering oral vaccines has the potential to reduce costs, simplify vaccine delivery and storage, as well as provide animal welfare benefits. Recombinant oral vaccines produced by plants, bacteria or yeast have been under investigation for their ability to induce the correct immune response. This project will investigate the microalgae C. reinhardtii as a host for recombinant oral vaccine production.
Constructs containing the SAV antigen will be produced at UCL using established techniques for genetically engineering the algal chloroplast. In order to enhance the process economics, we will investigate the influences of different lights and different feeding strategies. Following this, we will investigate how different harvesting and drying techniques may influence the yield and stability of the vaccine. The algal vaccine will then be tested in animal trials to establish how effective this may be in comparison to the established injection method of vaccination. The effectiveness of the oral vaccine will depend on the digestion and immune response elicted by the microalgae within the gut of the fish. Following the scale up of cultivation and the resulting animal trials, we will perform an economic assessment to understand the commercial viability of algal-based oral vaccines.

Technical Summary

In this proof-of-concept project we will aim to generate a transgenic line of C. reinhardtii expressing the E2 structural protein of Salmonid Alpha Virus (SAV). Via laboratory trials and technoeconomic analysis we will establish the effiacy and performance of dried algal biomass as an effective oral vaccine delivery system.

In order to produce the SAV antigen in the algal chloroplast, a synthetic gene will be designed to the E2 structural protein from SAV sub-type 3 (SAV3). This will be placed under the control of a strong constitutive promoter. Selection is based on the restoration of phototrophic growth by the wild-type copy of psbH carried on the plasmid.

Small scale cultivation of the recombinant C.rheinhardtii in ALGEM photobioreactors will be used to investigate the impact of light and nutrients on biomass productivity and product titre. This will then be scaled up using a modular hanging bag cultivation system. The process flowsheet for DSP will then be developed. Cell harvesting and dewatering via flocculation or centrifugation will be compared as a method for preparing biomass ahead of drying. Spray and freeze drying will be compared to find the most suitable method of formulating the oral vaccine, and examine the sensitivity of the SAV vaccine to heat and shear stress.

In the first part of the animal trial work we will assess the following:
1. Immunogenicity of recombinant algae vs. a validated SAV vaccine (AquaVac PD3).
2. Immunogenicity of algae with empty vector (adjuvant effect).
3. Relative percent survival (RPSend) provided by algal expressed SAV antigen delivered by injection.
In the second part of the animal trial work we will assess:
1.The fate of orally delivered recombinant algal vaccine antigen in the fish
2.Whether orally delivered algal vaccine can induce an immune response
3. The ability of orally delivered recombinant algae to protect salmon after viral challenge.

Planned Impact

The project will have impact in each of the areas defined by the BBSRC/NERC and RCUK: economy, society, knowledge and people.
In terms of economic impact the PI and CoIs have experience of working with industry, and the commercialisation of research findings and creation of spin-out companies. The constructs and data resulting from this project will be made available (after any IP protection) to the wider academic and industrial community via publication, and dissemination at conferences and meetings for example the BBSRC NIBB Networks. Furthermore, the method of producing and scaling up vaccine constructs can be made available to others for testing.
Beneficiaries of this work therefore extend beyond the immediate participants. It will benefit the UK economy by exploring new methods of manufacturing using algal synthetic biology and demonstrate the potential for oral vaccine delivery in aquaculture. The implementation of new/improved methods for cultivation and cell harvest of microalgae will benefit both the academic and industrial biotechnology communities seeking to translate research from bench to product.
The impact of knowledge generation from the research will arise from the communication and publication of new scientific advances and public outreach activities, detailed in the pathways to impact. In a wider sense, new methods for manufacturing with microalgae may be applied to other areas in industrial biotechnology and healthcare, resulting in gains in sustainabilty, in particular as part of a circular economy.
The societal impact will arise from improved animal health and reduced cost of vaccines which may enable cost savings attained through improved productivity to be passed to the consumer.
The impact on people will arise from training and career development of the PDRA and provision of skilled scientist in the industrial biotechnology sector. It will also enable an early career researcher to gain valuable PI experience with support and mentorship of more established academic CoI.
Should the oral SAV vaccine is shown to be effective, the impact on the UK could be the expansion of the salmon farming industry / the reduction of losses within the salmon aquaculture industry, therefore improving profitability and potentially enabling expansion of this sector. Demonstraitng the potential for algal-based manufacturing could lead to the development and growth of a new platform for vaccine production, and the development of brand new industries.
The findings will also influence future research directions, since the successful completion of this proposal will provide the foundations for further exploration in the area of algal synthetic biology. This will nurture a growing interest from other animal health and therapeutics industries who are interested in novel GRAS delivery systems.
We have established links with Government offices, trade organisations and business support bodies (e.g. Innovate UK), Societies (e.g. Society for Applied Microbiology, British Phycological Society) and algal associations (e.g. European Algal Biomass Association), and so are able to contribute positively to the framing of legislation regarding the control and exploitation of algal synthetic biology. Lobbying and involvement in the drafting of roadmaps and policy documents are already a key activity for both the PI and Co-I and this will continue under this project.

Publications

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Stoffels L (2019) Downstream Processing of TN72 for Recombinant Protein Recovery. in Frontiers in bioengineering and biotechnology

 
Description We have shown that it is possible to make different versions of a recombinant vaccine against salmon alphavirus using algal synthetic biology. We have expressed several versions of a vaccine in the algal chloroplast and measured their expression. The two most promising candidates have been scaled up and used for a fish trial to understand if immunity can be conferred through an oral vaccine, as opposed to injection. We have demonstrated that spray drying can be used as an alternative to freeze drying in order to prepare oral vaccines, and this will be important for future work in order to reduce cost and complexity of manufacture.
Exploitation Route The results of the process scale up work are of value to the academic and aquaculture sectors as we have demonstrated a manufacturing route for oral vaccines or other edible therapeutics that uses spray drying as a more simple technique of preparing biomass. This will be of value to the algal industrial biotechnology sector who are trying to develop downstream processes.
Pending the outcomes of the fish vaccine trials, we will be able to demonstrate the efficacy of an oral vaccination route. The results from this will inform future work on palatability of algal products that can be incorporated into fish feed. It will also provide valuable data on the potential of an oral vaccine through immunology tests.
Sectors Agriculture, Food and Drink,Healthcare,Manufacturing, including Industrial Biotechology,Pharmaceuticals and Medical Biotechnology

 
Description CEFAS Fish trials on algal vaccines 
Organisation Centre For Environment, Fisheries And Aquaculture Science
Country United Kingdom 
Sector Public 
PI Contribution We have used UCL facilities and expertise in algal biotechnology and bioprocess engineering to generate material for animal challenge trails.
Collaborator Contribution CEFAS have brought expertise in running aquaculture challenge trials, immunology and fish husbandry techniques. We have been able to avail of world-class aquaculture facilities in Weymouth in order to test the efficacy of an oral vaccine due to the involvement of CEFAS.
Impact This collaboration has resulted in an animal trial to test the ability of an oral vaccine to confer immunity to salmon alphavirus.
Start Year 2019
 
Title Edible vaccine against salmon alphavirus 
Description The product is an edible vaccine: briefly, a strain of Chlamydomonas reinhardtii which has been engineered to express various envelope proteins of the salmon alphavirus. We have scaled up the manufacturing process to make this vaccine. This has been tested in an animal challenge trial with salmon, to understand if the oral vaccine can confer immunity. Immunology results from the trial are currently being investigated. 
Type Therapeutic Intervention - Vaccines
Current Stage Of Development Initial development
Year Development Stage Completed 2020
Development Status Actively seeking support
Impact Understanding the scale up of the manufacturing process will help with other whole-cell therapeutics made in C.reinhardtii hosts e.g. endolysin proteins for antimicrobial activity. 
 
Description Talk at the Royal Academy of Arts 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact I presented at an event called "Rethinking Algae for Design" as part of the Algae Platform residency at the Royal Academy of Arts to accompany the Eco-VIsionaries exhibition. Since this event I have had several emails from artists, architects and designers who would like to collaborate on new projects related to / using algae. This has already lead to one new collaboration with the Design for Performance and Interaction course at the Bartlett. I have had one media enquiry from a journalist. I have received one invitation to the Algae Summit hosted by Atelier Luma in Arles.
Year(s) Of Engagement Activity 2020
URL https://www.royalacademy.org.uk/event/algae-design-eco-visionaries-architecture-talk