Fish and seafood are important protein sources for human nutrition. However, fishing yields cannot be increased at will and one third of commercially exploited fish stocks are already overfished. This development has led to a rapid upturn in global aquaculture which now already accounts for more than half of the aquatic food supply to markets around the world. This is a significant achievement, but it is linked to profound changes in aquaculture practices. Traditional farming methods are increasingly being replaced by highly productive technologies that are geared to the rapid growth of the fish, and this often requires deep interventions in their life cycle. These developments in aquaculture can be compared to those in agriculture in the 1950s. Like the “green revolution” in agriculture, the “blue revolution” in aquaculture is having significant environmental and social impacts. Both agriculture and aquaculture use intensive monoculture systems in which individual species are produced in limited habitats with high densities, often leading to problems with diseases and parasites, pesticide and chemical use, and locally posing a threat to ecological balance and the genetic diversity of individual wild populations. In the controversial public debates about the benefits and risks, advantages and disadvantages of biotechnology, the focus is usually on genetic engineering. Since November 2015 at the latest – that was when the US Food and Drug Administration (FDA) granted approval to the genetically modified Aqua Advantage Salmon from Aqua Bounty which grows to market size in one and a half instead of three years – many people have seen their fears confirmed that the last inhibition thresholds have now fallen and genetically modified organisms (GMO) will soon dominate the markets. Critics of aquaculture paint a distorted picture of this industry, accusing it of purely profit-oriented decision-making and too willing acceptance of possible risks to the environment and consumer health. Apart from the fact that this assumption isn’t true they deliberately ignore the advantages that transgenic fish would have for supplying markets and meeting the protein needs of the growing world population. Faster growth and higher resistance to disease, more usable muscle meat and greater temperature tolerance are just a few of the most significant advantages. Biotechnological processes help increase fi sh production while at the same time reducing the environmental and economic costs of aquaculture. What until recently was a topic for research is now often already being used in practice. Th e applications enabled by biotechnology range from eff ective breeding methods that enable rapid genetic advances, through the development of alternative feeds, to highly successful diagnostic and therapeutic methods that open up fascinating possibilities for preventing and combating diseases. And we are still only at the beginning of these developments which will probably revolutionise aquaculture in the foreseeable future. Th e changes not only aff ect individual areas but also, and more generally, the way in which fi sh and seafood will be produced in the future.
Gene editing simplifies genetic interventions
New findings have in recent times considerably expanded the spectrum of biotechnological methods. For example, alternatives have been found for certain methods that are rejected by parts of the public, and they will now likely have a greater chance of acceptance. Instead of the often contested gene transfer (in which genetic material is transferred from one organism to another in order to overcome species barriers that nature created over very large time scales in the course of evolution) it would in many cases be possible to use gene editing technology. This genetic engineering strategy also intervenes in the genetic material by modifying the “genome” of the organism but in contrast to gene transfer it usually only manipulates the species-specific genome. Special techniques are used to delete or replace sections of a gene. Among the currently particularly promising gene editing technologies for aquaculture one procedure, abbreviated to CRISPR / Cas9, stands out. The CRISPR / Cas9 procedure represents a kind of “gene scissors” with which the hereditary molecule DNA in the fertilized fish egg can be cut in a specific place (usually within the desired target gene). The cell recognizes the inflicted injury and repairs the damage independently. Since after the repair the DNA sequence often differs from the original version this method can be used to disrupt or prevent the expression of functional proteins. In contrast to gene transfer, gene editing does not involve the transfer of foreign DNA from other organisms and its integration into the target genome, i.e. it does not produce a transgenic organism. This method could therefore stand a better chance of being accepted by consumers. CRISPR / Cas9 technology has already been applied successfully to some fish species in aquaculture, e.g. Atlantic salmon, rainbow trout, catfish, tilapia and carp. However, a prerequisite for improvements in production characteristics is that they are significantly regulated by a single gene. For example, the elimination of the dead-end gene (dnd) makes the fish sterile and by modifying the myostatin gene (mstn), muscle growth can be increased. The content of the omega 3 fatty acid EPA in the fillet can be increased via the fatty acid elongase gene (elovl2). However, some of these mechanisms are not yet completely understood and the need for research remains high. In fish whose mstn gene has been switched off the entire immune system is often impaired, which makes them more susceptible to disease. Disorders of the elvol2 gene alter fat metabolism. Despite these problems, however, gene editing has such convincing advantages that its broad practical application – particularly in breeding – is definitely worthwhile. On the one hand, by switching off specific genes it is easier to identify which phenotypic characteristics are influenced and controlled by them. The more precise our knowledge of the relationships and dependencies between genotype and phenotype is, the greater the chances of success in breeding performance-enhanced strains and breeding lines. On the other hand, gene editing can also be used for precision breeding, for example to specifically implement the necessary genetic sequences in fish populations that lack certain desirable traits. This is done without “genetic dilution” of other important phenotypic traits, as is hardly avoidable with conventional breeding methods. Some methods are already proving themselves in practice Basically, aquaculture animals are particularly well suited to biotechnological processes because they are usually very fertile, i.e. they produce large quantities of gametes (eggs and sperm), fertilisation usually takes place outside the body, and the off spring can be reared under “artificial” conditions (“in vitro”). This opens up some worthwhile approaches for numerous biotechnological strategies such as the use of sex-reversal hormones to control reproduction or to make animals sterile, improve their growth, increase environmental tolerance, or suppress certain behavioural expressions such as aggressiveness. Biotechnological developments already used in aquaculture include synthetic hormones that are used, among other things, for the induction of sexual maturity, for the production of “mono sex stocks”, and for polyploidy breeding. Gonadotropin Releasing Hormone (GnRH) in particular has proven to be a universal biotechnological tool in fish farming. GnRH is a key hormone that initiates the cascadelike release of further hormones (e.g. LH and FSH) from the pituitary gland in fish and other vertebrates. Of the more than a dozen structurally clarified GnRH variants, about two thirds have been isolated from fish species. The best known of these is the salmon GnRH analogue which is used worldwide in commercial fish farming under the name “Ovaprim”. Without this hormone many fish species in aquaculture could not be made ready for spawning and reproduced. Biotechnological methods are also used for some “genome manipulations” (“chromosome engineering”) such as the production of tri- and tetraploid animals and gyno- and androgenesis. The classical methods for manipulating chromosome sets (polyploidization) use thermal processes (cold or heat shocks), hydrostatic pressure or chemical treatments (e.g. colchicine, cytochalasin B) to produce tri- or tetraploid animals. Triploids can also be produced by crossing tetraploids with normal diploid organisms. Triploid fish are sterile and put the feed energy into growth rather than gonad maturation and reproduction. This works not only with fish but also with invertebrates such as oysters. Triploid Pacific oysters are sterile and grow 15 to 150% better than comparable diploid oysters. During gynogenesis, the male genome of the sperm is inactivated so that the development of the animals is controlled exclusively by the maternal heritage. In aquaculture, it can also be useful to influence the sex of the animals held if only one of the sexes, i.e. either male or female, possesses the economically desirable characteristics. Monosex populations can be created by hormonal sex reversal with sexual steroids in the early stages before sex differentiation begins or by genetic methods. Monosex is common practice for tilapia, for example, because male animals grow faster and to larger sizes than females. Apart from that, tilapia held in mixed populations mature very early, hardly grow at all and reproduce uncontrollably. In the case of salmon and sturgeon, females are preferred because they grow better and provide the coveted caviar. Precision breeding moves into the realm of the feasible Classical selection programmes will continue to be the decisive driving force for breeding improvements worldwide but biotechnological methods are becoming more and more important for both short- and long-term strategies. Compared to farm animals, domestication in fish is not yet very far advanced, and there is considerable potential for genetic improvement in commercially important species. Breeding programmes exist at different levels, for example, for Atlantic salmon, carp, gilthead seabream, hybrid striped bass, tilapia, sea bass and rohu carp. The focus is usually on growth rate which can be improved by up to 20% per generation in particularly good breeding lines. Disease and stress resistance, late onset of maturity, and meat quality are also important selection targets. In the case of short-term strategies for genetic improvements biotechnological processes enable rapid progress. Genome mapping, which shows the relative positions of genes in the chromosome, and DNA markers facilitate the selection of those animals that possess the desired traits and are therefore particularly suitable for breeding. Marker-assisted selection (MAS), for example, is already used for rainbow trout. Genetic markers can also be used to identify individuals and family groups and keep them in common groups which greatly simplifies breeding programmes. For certain purposes, especially long-term breeding programmes, the long-term preservation of biological material can be important. This is usually done in liquid nitrogen at -196°C (“cryopreservation”). Cryopreserved spermatozoa may not yet be as important in the fish sector as in agriculture, but there is certainly potential for the use of this biotechnology which could facilitate selective breeding. Especially since this would also solve the problem that males almost always mature before females. Cryopreservation also enables the preservation of valuable genomes, which will in the future make it possible to set up gene banks for aquaculture. Molecular genetic diagnoses, genetic markers and DNA fingerprints also benefit consumers because they are highly sensitive and very precise which improves the accuracy of product traceability. Incorrect labelling, species substitution, undesirable admixtures and other cases of food fraud are identified quickly and without doubt. And not only for whole fish, but also for fillets, frozen products, eggs and larvae or canned products.
Optimized feed and effective health protection
Important applications of biotechnology in aquaculture include fish nutrition and feed. The importance of biotechnological processes is growing in line with the replacement of fish meal and fish oil in aquatic feeds by alternative raw materials, which often require complex preparation and “pre-processing” before they can be used for fish nutrition purposes. This applies in particular to plant material that is broken down using biotechnological methods in order to reduce the content of harmful or nutritionally inhibiting substances. A worthwhile but complex method is to breed plants that have fewer nutritional inhibiting factors and an amino acid profile that is more suited to fish. Genetically modified yeasts are also produced for fish nutrition. These yeasts contain, for example, significantly more carotenoid pigments, which are indispensable in the breeding of some species. The addition of exogenous digestive enzymes to fish feed has not yet produced the hoped-for benefits. The use of so-called probiotics seems much more promising. They are increasingly replacing antibiotics and they improve the external microbial environment in ponds and other aquaculture systems. An example of the successful use of probiotics in feed is shrimp cultures where they effectively prevent or inhibit some viral diseases of shrimps. In the case of viral diseases prevention of the pathogen is crucial. Biotechnological tools such as gene probes and polymerase chain reaction (PCR) enable rapid and reliable detection of such pathogens. The enormous increase in aquaculture production of whiteleg shrimp (Litopenaeus vannamei) is largely due to the use of specifically pathogen-free (SPF) and specifically pathogen-resistant (SPR) shrimp on farms. In addition to molecular diagnostic detection methods and immunostimulants, vaccines that protect fish and shellfish from dangerous diseases are increasingly used in the field of fish health. Researchers around the world are working on a new generation of highly effective vaccines that no longer consist of “inactivated pathogens” but of genetically modified microorganisms, protein subunits and DNA fragments. But even with that the possible applications of biotechnological and microbial methods in aquaculture are far from exhausted. Microbial communities can, for example, optimise biofiltration and detritus recycling, the nutrient and energy cycles in waters and thus sustainably support and increase productivity in aquaculture. Biotechnology also has much to offer in the field of “natural product research”, which promises commercially valuable pharmaceutical products such as pigments, oils, sterols, alginates and agarose from micro- and macroalgae. Many of these developments are still in their infancy but the wealth of species (which is many times greater in the oceans than on land) already raises enormous hopes. Only recently, researchers isolated compounds from the blood plasma and tissue extracts of horseshoe crabs (Limulus polyphemus) that have been shown in laboratory tests to have the potential to inhibit and destroy human colon cancer cells and malignant melanoma cells.