• 21
  • FEB
  • 2018

Why are “fish oils”, or omega-3 long chain polyunsaturated fatty acids (LC-PUFAs), so important?
The omega-3 LC-PUFAs, EPA (eicosapentaenoic acid) and DHA (docosahexaenoic acid), are important for human health and nutrition. They modulate both metabolic and immune processes, and they confer health benefits in areas of coronary heart disease (CHD) and neurodevelopment (Nutrition Reviews Vol.71(10):692-707). They are found in marine fish and algae, as noted in the associated article, "GM Field Trial Planned".

Strong evidence shows that consumption of fish, and oily fish in particular, lowers the risk of death from CHD by 36% (Joint FAO/WHO Expert Consultation on the Risks and Benefits of Fish Consumption, 2011 ).

National and international guidelines for the general population recommend consumption of at least 250 milligrams a day of omega-3 LC-PUFAs or at least two servings a week of oily fish for optimal protection against CHD ( J Am Coll Cardiol, 2011, 58: 2047-67).

What is the natural source of EPA and DHA, and of the ketocarotenoid pigment, astaxanthin?
As with humans, most fish obtain EPA, DHA and astaxanthin through their diets. Thus, “fish oils” are not actually made by fish, neither is their pinkish pigmentation; they are synthesised by marine microbes (algae), which form the base of aquatic food webs.

Farmed fish receive EPA and DHA through their feed; if not, the resulting fillets lack these important fatty acids. Fish farms consumed around 80 per cent of all fish oil harvested from the oceans in 2011 (as fish), and this ratio has not changed significantly since then.

As the human population increases and fish stocks in the oceans diminish, fish farming (aquaculture) has become a major source of fish for human consumption. EU aquaculture is valued at €3.2 billion (£2.8 billion) and accounts for one-quarter of the EU’s production of fish, molluscs and crustaceans. According to Scotland Food and Drink, the export value of Scottish salmon in 2016 was £600 million, and Scottish aquaculture contributes over £1.8 billion annually to the UK economy.

Aquaculture provides high-quality protein for human nutrition, and more efficiently than terrestrial livestock farming, according to Skretting, an Australian manufacturer and supplier of aquafeed, but the industry is heavily dependent on marine-derived fish oil and fish meal.

Fish oil and meal are usually sourced through the harvesting of “feed-grade” species, which would not normally be suitable for direct human consumption. However, these so-called “reduction fisheries” are at their sustainable limits, and another source of omega-3 LC-PUFAs is required.

Why is this experiment necessary?
We have already successfully developed Camelina plants, specifically Camelina sativa, in our laboratories and glasshouses to produce fish oils in their seeds, tested them in the field in 2014 and 2015, and published the results (Usher et al., 2015; Usher et al., 2017).

We have now further developed our Camelina plants to produce more fish oil, and we wish to test these new plants under “real-life” conditions in the field.

Why do you use GM technology?
Current plant sources of omega-3 PUFAs, such as flax seed, do not produce the long chain varieties, EPA and DHA; instead they produce shorter chain omega-3 PUFAs, such as ALA (alpha-linolenic acid). ALA does not confer the beneficial health properties associated with EPA and DHA.

There is no economically viable or sustainable source of wax esters of the type we wish to make (similar to those found in Spermaceti whale oil); regarding astaxanthin, only very few plants accumulate this ketocarotenoid pigment, usually in their flowers. Among them is the Adonis plant from which genes were copied to make the pigment in our Camelina plants.

Could algae produce omega-3 LC-PUFAs and astaxanthin without genetic modification?
They could, but it would require vast amounts of algae and a quantum leap in existing technology; making this route economically viable has proved exceptionally challenging for the algal sector.

At Rothamsted, we explore all possibilities for the development of sustainable sources of EPA and DHA, including research on algae and diatoms. Although we have recently demonstrated significant progress in this area, we are still at earlier stages in this work than we are with our work in Camelina plants. Producing these oils in Camelina requires the use of only standard, established farming practices and machinery.

What is the nature of the genetic modifications you have made?
We synthesised the gene sequences that encode the enzymes involved in the production of omega-3 LC-PUFAs, astaxanthin and wax esters, and optimised them to work in Camelina plants. We did the same to improve the plant’s photosynthetic capability with bigger leaves; we did not need to synthesise any genes to increase its stem thickness.

For the omega-3 EPA and DHA, the genes are found in photosynthetic marine organisms, such as phytoplankton and other lower eukaryote species. For the astaxanthin, the genes are found in the Adonis plant. For the wax esters, the genes are found in marine bacteria and higher plants.

For enhancing photosynthetic capability, with bigger leaves, the genes come from the bacterium Escherichia coli; and for the other architectural trait, thicker stems, we use an Arabidopsis gene.

We have produced fourteen varieties of plants: one contains eight synthetic genes; one contains seven synthetic genes; three contain six synthetic genes; one contains five synthetic genes; and two contain four synthetic genes. These more complex metabolic engineering lines are designed to produce the omega-3 fish oils.

We also have two varieties of plants that contain three synthetic genes, for the production of astaxanthin and wax esters. The remaining GM plants contain only one gene, for the architectural traits (bigger leaves and thicker stems) and for a wax ester line.

We needed to introduce this number of synthetic genes because the production of omega-3 LC-PUFAs, astaxanthin and wax esters requires processes that involve many steps. To maximise production, we had to help the plant’s biosynthesising machinery to shift from making ALA to making EPA and DHA, or to produce astaxanthin and wax esters.

All the genes producing LC-PUFAs, astaxanthin and wax esters are active only in the seeds of the Camelina plant; and they are most active during the mid-stage of seed development. Gene expression has not been observed in any other vegetative tissue during the life cycle of the plant.

All plants expressing the inserted genes look the same as normal plants (the control plants) in the glasshouse. The inheritance of the inserted genes over five generations follows normal rules of Mendelian genetics. No difference has been observed in seed set, seed size or germination. No difference was observed in fertility. The vegetative performance of the transgenic plants was unaltered.

In the case of the architectural traits, these genes are expressed in all parts of the Camelina plant; as expected, they cause some modification to the morphology of the plant, including either thicker stems or bigger leaves.

How important is this research?
This research is very important at three levels: scientifically, the delivery of a plant that produces seeds with the above characteristics depends on employing the most complex genetic and metabolic engineering approaches used in plants to date; economically, we are close to delivering plants able to produce seeds enriched with commercially valuable omega-3 LC-PUFAs; and socially, the research offers societal benefits because of its potential impacts on both human health and the environment.

How have you communicated this research and the possibility of a field trial?
We have been working on this area of research for more than 15 years and information has been made accessible and available on our website over this period. We have also published an assortment of material, from fact sheets to press releases, and spoken about this work at public and scientific meetings, and to the media, on many occasions. The results of our academic research on this topic have been published in open-access, peer-reviewed journals.

Will independent experts and the public be consulted?
As part of the statutory process co-ordinated by the Department for Environment, Food & Rural Affairs (Defra), there is a period of public consultation before a decision on whether the field trial can go ahead. People can make representations to Defra on any environmental risks that they think the trial might pose. These concerns will be considered by the independent experts of ACRE, the Advisory Committee on Releases to the Environment.

ACRE is a statutory advisory committee appointed under section 124 of the Environmental Protection Act 1990 to provide advice to government regarding the risks to human health and the environment from the release of genetically modified organisms (GMOs).

Why is there a need to carry out this research in the UK?
The UK has world-class plant scientists who should be at the forefront of developing scientific and technological advances that can make agriculture more efficient and sustainable. Investment in biotechnology promotes the UK’s competitiveness and ensures long-term benefits to taxpayers.

Are fish feeding experiments planned with any oil obtained from this field trial?
No, fish feeding experiments are not planned as part of this field trial. The seeds collected will be used only for analysis of oil content in the laboratory. However, we are planning to extract oil from the seeds of plants grown in the glasshouses for fish feeding experiments.

Will the trial be safe?
Yes. Rigorous regulations govern the planting of GM crops in the UK, overseen by Defra and its independent advisory committee, ACRE (see above). There will also be inspections during the trial, carried out by the Genetic Modification Inspectorate, which is part of the UK’s Animal and Plant Health Agency. The ensuing inspection reports are made publicly available.

At Rothamsted, we also have an internal Biosafety Committee, which makes risk assessments of proposed field trials before applications are submitted.

What is the process of approval for research using GMOs?
At Rothamsted, as at many universities and research institutes around the world, work is taking place using GM plants and microbes for a variety of projects and to address specific scientific questions.

Research is conducted in our laboratories, in our glasshouses and, in some instances, in the fields of our research farm. Where research using GMOs is undertaken, relevant regulations are followed and risk assessments carried out.

Rothamsted’s activities are compliant with the law and our Biosafety Committee oversees all projects that involve GMOs. There are several key pieces of legislation specifically concerned with the contained use of GMOs.

The main piece of legislation, covering both human health and environmental aspects of work in laboratories and glasshouses with GMOs, is the Genetically Modified Organisms (Contained Use) Regulations 2000, as amended.

In risk assessments of our procedures, we follow best practice as specified by the Compendium of Guidance, which has been put in place by the Health and Safety Executive (HSE), in conjunction with Defra and the Scottish Government.

Occasionally, a research project requires experimentation using GMOs in the field, for which approval from Defra is required. We have to carry out a full risk assessment of the project, which is scrutinised by the institute’s own Biosafety Committee, and submit an application to Defra.

In the application, we describe the nature of our experiment, the type of plant material and the specific genetic modifications that we have carried out. We are also required to assess the risks to human health and the environment.

The application becomes publicly available on the Defra website and our application is independently assessed by ACRE (see above). As soon as the application is submitted, there are 48 days for public consultation, carried out by Defra; any member of the public can comment or ask any relevant question.

Once the review of the risk assessment has been carried out, ACRE make their recommendation to Defra, which considers it along with any public representations that the department has received. Defra then decides whether to grant a consent to conduct an experimental field trial (with or without specific conditions).

What security measures will you be taking?
Fences will surround the experimental sites to prevent the entry of rabbits and other large mammals. There is also CCTV.

Why are these measures necessary?
This trial will be a controlled experiment and we want to ensure that the experiment is conducted with the scientific rigour and high standards expected of Rothamsted. To do so, we need to ensure that rabbits, dogs, other large animals and people do not wander into the field and interfere with the experiment.

What about the risks of cross-pollination with local wild plants?
It is highly unlikely that any of the modified genes would transfer into other crops. While Camelina sativa can intercross with other members of the Camelina genus, in particular Camelina microcarpa and Camelina alyssum, these weeds are not found anywhere on the Rothamsted farm sites where the trials would take place.

Within the larger Camelineae tribe, species such asArabidopsis lyrata, Capsella bursa pastoris and Neslia paniculata seem not to cross with C. sativa or, if they do, viable seeds do not result (Julie-Galau et al., 2013).

No cross-pollination, either natural or forced, has been observed betweenC. sativa and members of the Brassica genus, such asB. napus (oilseed rape), B. juncea (brown mustard), B. rapa (field mustard) and B. nigra (black mustard).

In the laboratory, artificial hybridisation between protoplasts (plant cells without cell walls) of C. sativa and B. napus, B. carinata and B. oleracea has been reported, but with low success and/or sterile hybrids (see, The Biology of Camelina sativa (L.) Crantz/Camelina)

Although cross-pollination is highly unlikely, management measures will still be in place to ensure that such events do not occur. There will be no sexually compatible species grown within 1000 metres from the boundary of the site and no sexually-compatible wild relatives of C. sativa have been found in the vicinity.

The trial’s perimeter will have a border of non-GM C. sativa, at least 7m wide, to function as a pollen barrier. The machinery for sowing seeds will be filled on the trial area, and it will be thoroughly cleaned before leaving the trial area. To minimise the possibility of seed loss, the plants will be harvested just prior to full maturity. All straw will be chopped and left on site.

Could gene transfer affect farming produce nearby?
The chances of gene transfer affecting farming produce if this experiment goes ahead are exceptionally low (see above).

Could cross-pollination enable food crops to produce wax esters that are said to cause digestive problems?
Wax esters are common on land and in the sea, and there is a greater chance of cross-pollination from existing sources that from GM camelina plants. For many people, wax esters form part of a nutritious diet, as they are naturally present (as part of the cuticle) in most plants and vegetables.

Could butterflies be damaged by eating fish oils produced by the GM Camelina plants?
A study by Stefanie Hixson and others, published in 2016, concluded that “the presence of EPA and DHA in diets of larval P. rapae may alter adult mass and wing morphology; therefore, further research on the environmental impacts of EPA and DHA production on terrestrial biota is advisable.”

Pieris rapae is the cabbage white butterfly, a crop pest that, in its native environment, feeds on leaves of the cabbage family, not on EPA or DHA. The Hixson specimens were fed in laboratory conditions with experimental diets.

The artificial diets were provided to the larvae as a formulation; EPA and DHA were isolated from algae directly and added in the feed as free fatty acids. By contrast, the EPA and DHA synthesised by the GM Camelina plants are a component of the oil synthesised in seed. Cabbage white butterflies do not feed on seeds in their native environment at any stage in their development.

Rothamsted has analysed leaf tissue of GM and non-GM Camelina plants; the institute found that the leaves have identical fatty acid profiles and are devoid of EPA and DHA. These data will be submitted for peer review as part of a broader study later this year.

Are you going to test the safety of the enriched oil produced?
The aim of the proposed trial is to test only the performance of the plant under field conditions and its ability to produce omega-3 fish oils and the other compounds and traits, as has been observed in glasshouse experiments.

How is this Camelina better for the environment?
We hope the field trial will help to tell us whether we can use plants to produce a more sustainable source of healthy fish oils. Internationally, the aquaculture industry is working hard to increase the sustainability of fish feed production practices.

Do you foresee any environmental problems from Camelina?
No. C. sativa originated in Europe, and was historically grown across south-eastern Europe and south-western Asia. It is a native species in many European countries, including the UK. In recent years,C. sativa has not been widely cultivated as a crop in the UK. C. sativa is grown as a crop in Canada and parts of the USA.

What are the effects on biodiversity of this research?
This is a small-scale, highly controlled experiment over the course of just three years. We do not anticipate any effects on biodiversity.

How did you make the GM plants?
We synthesised gene sequences encoding enzymes involved, for example, in the production of omega-3 LC-PUFAs and other compounds, and optimised them to work in Camelina plants (see above). We developed “gene switches” (or “promoters”) from other genetic material to determine when and where the synthetic sequences work. The switches come from genes in other plants, such as Brassica napus, Arabidopsis and Linum usitatissimum and, in one instance, from the Cassava vein mosaic virus.

We have also developed genetic “end-signal” sequences to co-ordinate the performances of the genes that we have inserted into the plant. The material for these sequences come from various organisms, including Arabidopsis, Agrobacterium tumefaciens and Cauliflower mosaic virus.

Will your experiment be legal?
Yes. GM experiments are allowed in Europe if strict regulations are followed. This GM field trial will proceed only if it is authorised by Defra (see above).

Will the public be able to see the trial?
Yes. With Rothamsted’s permission, it will be possible to see the trial.

How big is the trial?
The area of the proposed trial at Rothamsted, including controls and spacing between GM plots, will cover 2130 square metres, including the pollen border; that at Brooms Barn covers around 6000 square metres.

Who will be paying for the trial?
Rothamsted will pay for this trial from the strategic funding that it receives from the Biotechnology and Biological Sciences Research Council (BBSRC), specifically as part of Tailoring Plant Metabolism, one of the institute’s five strategic programmes (2017-2022). The institute is also supported by the Lawes Agricultural Trust (LAT).

Isn’t this trial just a waste of taxpayers’ money?
No, on the contrary. For many years, Rothamsted has done this research in laboratories and glasshouses; over the past four years, the institute has taken the work into fields (with Defra’s approvals). Published results from field trials conducted in 2014 and 2015 demonstrate that the plants can stably produce omega-3 fish oils in the “real world”.

We have now developed plants that have the potential to produce commercially-relevant yields of omega-3 fish oils. This experiment will assist in determining whether the technology could provide environmental and financial benefits for the UK economy and the UK taxpayer.

Have you received industry funding for this work?
No. Rothamsted does receive some funding from industry (around 10% of its income) and aspects of the development of this technology over the past 15 years have been made in collaboration with industrial partners, but this particular trial is publicly funded.

Our overarching philosophy is that we need to work with government policymakers, non- government organisations (NGOs), agribusinesses and farmers if we are to deliver the knowledge, innovation and new practices that are needed to increase crop productivity and quality and to develop environmentally sustainable solutions for agriculture and aquaculture.

Working with industry forms a crucial component of this philosophy if we are to turn our scientific knowledge into technologies that can benefit end-users and consumers; only industrial partners have the necessary infrastructure to develop and distribute innovative technologies to those who need it.

We recognise that the UK taxpayer is the main funder of our research, and we have sought the public’s input, as well as that of our stakeholders, in developing guiding principles of how we should work with industry.

Who owns the patents in this area, and who will own the results of this research?
Many patents have been filed over the years covering various aspects of this area of research, and they are owned by a number of organisations, including Rothamsted. The institute will license the technology under its terms and conditions to recover investment for the UK taxpayer.

How will you ensure that the technology’s rewards benefit society in general?
When licensing our technology to commercial producers, our primary goal is to ensure maximum benefit to society. We therefore take steps to ensure that licensees achieve this.

Is there a market for this product?
Yes. Fish oils are now significantly more valuable than vegetable oil; they are currently estimated to be worth over £5 billion per year globally.

Are you going to put the knowledge gained from this trial in the public domain?
Yes. After peer review, the results of this field trial will be published in the appropriate scientific journal and we will make them accessible on our website. We are very proud of our track record of open access publication and the dissemination of our results.