Example 2: Transgenic Fish

The modification of animals is very different from that of plants. Although we are able to regenerate animals directly from a single cell, the bombardment of a tissue as used in transforming plants is not available. The generation time of an animal (particularly large animals rather than insects) is too long to allow selection techniques which allow the transformation of a very few cells to provide a significant source of transgenic animals. For large animals the transformation must result in a very high success rate, probably more than 50%.

Although modification of animals is very different from that of plants, the questions which have to be considered in order for a risk assessment to be performed are very similar.

Techniques for transforming animals include microinjection, where the DNA is injected directly into a cell or the use of retroviruses which should result in stable integration of the genes within the genome of the transformed cells.

One group of animals which has proved controversial is fish, both because they may prove relatively easy to modify, and because they are not easily contained. Recommendations on marine releases made by the International Council for exploration of the Sea (ICES) are published in the "1994 Code of Practice on the Introductions and Transfers of Marine Organisms". The discussion which follows is largely based on a publication by the British Department of the Environment entitled "Guidance for the experimental release of Genetically modified fish" referenced in footnote .

If fish are modified for aquaculture the likelihood of escape into natural ecosystems is thought to be high, and the fish might well carry traits which might result in significant impacts on ecosystems. In 1998 there had been experiments on 31 species using approximately 40 different DNA constructs. Most of the work has been on

• growth enhancement,
• environmental tolerance, including tolerance to extremes of temperature, salinity, heavy metals or crowding,
• reproduction, where inhibition of early maturation would reduce loss in aquaculture, and
• disease resistance, where there is potential for research on viral, bacterial and parasitic infections.

A major problem when compared to plants is the generation time for species of fish used in aquaculture -- generation times may range from less than one year for Tilapia to a few years for salmonids.

"Microinjection into the fertilised egg is the most commonly used technique for introduction of a DNA construct. Unlike mammalian eggs, it is generally not possible to visualise the pronuclei or nucleus of fish eggs, and microinjection is generally into the egg cytoplasm before the first cell division. This often leads to high levels of mosaicism, with integration of one or more copies of the introduced DNA often occurring after the first cell division". Other techniques which have been used include electroporation of eggs and sperm, or soaking of sperm in the DNA construct prior to fertilisation. Biolistic techniques have also been attempted.

The hazards which could be identified for which likelihood would have to be considered include knowledge of the molecular biology and physiology of fish. The basic science 'database' for fish is limited; although the potential for modification for use in aquaculture is vast, detailed knowledge particularly of the physiology of most fish species is limited

Infomation requirements include:

• capacity to survive, breed, establish and spread to other habitats

If the GM fish proposed for release has very specific habitat requirements the distribution of the fish after release may be able to be predicted, and monitored. If the fish roams over a wide variety of habitats, the capability to monitor will be greatly decreased. The impact and survival of the modified fish depends on their ecological fitness relative to wild stocks. The performance of these fish, however, could be very different from that observed in containment. The relative fitness could be unchanged, increased or decreased due to the modification. If decreased, the new form would be less likely to establish self-sustaining populations, but it could take many generations before the genetically modified fish disappeared.

The capacity to breed will be influenced by the ability of the fish to survive in the open environment but may also be dependent on the transgene or on the site of insertion of the gene into the genome. If the GM fish is capable of reproduction in the open environment, it may be able to spread to other habitats, establish viable populations, transfer genetic material and compete with other organisms.

• Behavioural changes in the modified fish and their descendants
• Physical and physiological changes in the modified fish

The genetic modification may cause physical or physiological changes. An introduction of a growth hormone to increase growth rate may require a greater food intake, or result in the modified fish eating smaller relatives. This would deplete the stocks of 'wild-type', even if the modified fish was less fit, or even sterile. A fish modified for increased tolerance to low temperatures may have an increased capacity to spread into a wider range of habitats. Phenotypic changes may provide the fish with a competitive advantage for food, shelter, mates and suitable breeding sites. Damaging competition with wild stocks has to be considered as a negative outcome of an introduction.

• Potential for and consequences of transfer of the inserted genetic material to other fish or organisms

Successful breeding between modified fish and wild relatives could result in a change of the genomes of the wild stocks. Problems which might arise would then depend on the inserted gene, the frequency of transfer and the fate of the offspring. The spread of the gene from modified fish to other species by hybridisations would be of concern if the resultant hybrid offspring carried an undesirable trait.

• Competition with other organisms
• Phenotypic and genotypic stability

Damage to the environment may be delayed and long term as it may be caused by the descendants of the originally released fish.

Once the hazards have been identified, the likelihood of these occurring must be estimated, and risk management procedures must be considered. A serious problem is that physical containment is rarely perfect -- Penman (reference ) suggests that the introduction of modified fish into aquaculture should be considered as an intentional introduction.

It may be possible to use biological containment, by (for example) ensuring sterility.