Brewers’ yeast as a supplement in aquaculture

by Jan Frericks, Leiber GmbH, Germany

The effect of stress caused by environmental pollution and farming conditions on the health and yield of fish in intensive aquacultures is becoming increasingly important.
Factors such as stocking density, contamination, toxins, pollutants and outbreaks of disease have a negative effect on the immune status of the fish. The consequence of this is an increased susceptibility to infection through bacterial, viral, fungal or parasitic pathogens. Increased loss rates and reduced growth performance result in lower profitability for the fish production industry.
As a consequence, the monetary and quantitative overheads for the vaccination and medicinal treatment of the fish increases. In many cases, antibiotics are given not only therapeutically, but also prophylactically as a standard additive in fish feed.
The intensive farming methods used for fish cultivation in aquatic environments with a direct connection to the groundwater are especially liable to facilitate the very rapid and direct spread of problematic production residues to humans. Resistant pathogens and germs do not just limit the effectiveness of therapeutic antibiotics for fish. The transfer of genes for resistance between different species of bacteria is accelerated, leading to an exacerbation of the problem of resistance in the treatment of human diseases worldwide.
Future-oriented production methods in the fish farming industry should therefore be targeted towards minimising the use of antibiotics and medicinal drugs. It is of great importance to analyse the negative effects caused by environmental pollution and farming methods.
More crucial still will be to influence the animal’s metabolism so that external toxins have a lesser impact, even under intensive conditions. A healthy gut and a functioning non-specific immune response are fundamental prerequisites for this.

Excellent for the gut
Brewers’ yeast cells are like miniature power houses, and are responsible for the alcoholic fermentation that takes place during the brewing stage of beer production. In the course of the fermentation of malt extract, high concentrations of minerals and trace elements, amino acids and nucleotides, B vitamins and enzymes, as well as many micro-nutrients accumulate within the cells of the yeast species Saccharomyces cerevisiae. Being organically bound ensures high availability of these active substances. Dried brewers’ yeast is used very often in fish nutrition due to the high bioavailability of the constituent compounds.
In addition to this, brewers’ yeast has cell walls that are composed of mannan-oligosaccharides (MOS). This complex network of mannans and ßglucans serves as a substrate for the beneficial gut flora. The fish’s limited digestive tract benefits in particular from the prebiotic properties of the yeast cell walls, which stabilise the gut and ensure a healthy balance of microflora (eubiosis). In addition to this, the mannan-oligosaccharides in brewers’ yeast are able to bind harmful toxins in the food, and thus inhibit their absorption and resultant metabolic harm. Last but not least, the formation of a biofilm on the intestinal mucosa enhances this protective barrier against pathogens.


Glucan and the immune system
The cell wall of brewers’ yeast comprises approximately 20-25 percent mannans and 25-30 percent ß-glucans. ß1,3/1,6(D)glucan molecules can be isolated from it using special hydrolytic processes. The molecules consist of characteristic (1,3)-beta-glycosidic linked D-glucose subunits connected with with irregular beta-(1,6)-linked side chains of various length. Only this free ß-glucan structure from Saccharomyces cerevisiae has an immunomodulatory effect on the metabolism.
In contrast with intact yeast cells or mannan-oligosaccharides, free ß1,3/1,6(D)glucan molecules are able to pass through the protective epithelial barrier in the gut with the help of specialised M cells. In gut-associated lymph tissue (GALT), ß-glucans act like antigens, stimulating specific macrophage receptors with their characteristic surface structures (epitopes) (Engstad and Robertsen 1993). A cascade of immune responses is triggered, and non-specific immune system cells such as monocytes, natural killer cells, B-cells, T-cells or lysozymes are released or activated. They put the animal on a high state of alert and preparedness to defend against all types of foreign attack. What differentiates this from an actual infection is that ß-glucan does not possess any pathogenic properties, and acts without causing any adverse health effects.
A quality criterion for the effectiveness of ß-glucan products is not only the source and characteristic molecular structure, but also the purity of the product. A minimum content of 70 percent pure ß1,3/1,6(D)glucan should be aimed for. The standard grade Leiber® BetaS has a ß-glucan content of 80 percent. The metabolic activity of this product has already been tested on many animal species and verified.
Tests were recently performed on several species of fish that confirm a stimulatory effect on the non-specific immune system. Rainbow trout and carp received 0.02 percent ßglucan (Leiber® BetaS) administered in their feed ration. The potential killing activity of phagocytes, the proliferative response of T and Blymphocytes and the concentration of immunoglobulins and lysozymes in the blood serum was measured four weeks and eight weeks after beginning administration. In two repetitions, a significant stimulation of these parameters was demonstrated in both rainbow trout and carp.
In subsequent infection studies using two bacterial (Aeromonas salmonicida; Yersinia ruckeri) and one viral pathogen (IPN virus), the survival rate of rainbow trout and carp with 0.02 percent Leiber® BetaS in their feed showed an absolute increase of 30-40 percent (Siwicki, et al. 2008; Siwicki, et al. 2009).


Oral administration tests
Purified ß1,3/1,6(D)glucans enhance the animals’ non-specific immune response. This is of particular value in the fish farming industry, as there are multiple stress and environmental factors that impact and stress the fish. If they become infected, the specific immune system is only able to respond slowly and inadequately. In such situations, a heightened non-specific capability can support or accelerate the specific immune response and the production of specific antibodies.
Fish are subject to similar, added stresses during vaccinations too. A study by Siwicki et al. (2011) investigated the effect of orally administered ß1,3/1,6(D)glucan (100mg or 200mg Leiber® BetaS per kg of feed) on the antibody secreting cells (ASC) and specific antibody titres after immunisation of rainbow trout fingerlings (Oncorhynchus mykiss) by immersion with anti-enteric redmouth disease vaccine (AquaVac ERM). Inoculation was performed one week after the start of administration with Leiber® BetaS.
These two parameters were measured on day seven, 14, 21, 28 and 40 in the blood serum and adrenal glands. Both dosage levels stimulated the number of specific ASCs and specific antibody levels, whereby 0.02 percent Leiber® BetaS in the fish feed was more effective. In each case the improvements were significant from day 21 onwards.
The beneficial effect of ß1,3/1,6(D)glucan is well known in the fish nutrition industry. The administration of 0.02 percent Leiber® BetaS in fish feed activates the non-specific immune status of the fish on the one hand, and on the other, acts as an adjuvant during vaccinations, thus enhancing the immunocompetence of the fish. Its safe and simple method of use, as well as the fact that ß-glucan from Saccharomyces cerevisiae is harmless to fish and the environment, will become yet more important in the future.     ■

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Ornamental fish and invertebrates for home aquaria

by Dr Jack M James, Aquabiotegch Group, Malta

The trade in ornamental fish and invertebrates is a truly global industry, generating many millions of dollars, and touching the lives of a vast range of people. From artisanal fishermen in Indonesia, to importers and exporters in Singapore and Spain, farmers in the Czech Republic and Florida and ending with the home aquarist in any one of hundreds of countries worldwide, the appeal of ornamanetals is worldwide.
While freshwater ornamental fish are largely farmed, thereby providing a sustainable and renewable supply, marine species are largely wild caught, leading to a potential for species loss, ecological imbalance, and habitat degradation. As the ornamental industry provides livelihoods in many places where there are very few opportunities for employment, it is important that the industry is encouraged to grow, but it is essential that proper monitoring is in place to ensure that this growth is sustainable, in terms of individual species, population ecology, and habitat preservation.


Industry scale
Livengood and Chapman (2008) estimated that some 1539 species of marine and freshwater fish, 102 species of hard and soft coral and 293 species of invertebrates were traded globally. According to FAO statistics from 2004, as summarised by Ploeg (2004), between 1974 and 2004, the number of countries reporting ornamental fish exports rose from 28 to 146, and this number is expected to continue to rise.
It is expected that most of these are developing countries which see the export of ornamental fish as a means to increase employment and generate wealth. The worldwide value of exports in 2004 was reported to be US$251m, a rise of US$230m in the preceding 30 years at an average 14 percent per annum, with a retail value of approximately US$2.2bn.


A further FAO report in 2008 valued exports at US$278m in 2005 (Livengood and Chapman, 2008). At these rates, it could be estimated that global exports now value over US$600m, although the effects of the global economic slowdown are not yet known for the sector.
In terms of the division of these exports between regions and countries, 55 percent of the 2004 exports came from Asia, while 25 percent came from Europe, mainly the Czech Republic. Between 1974 and 2004, the number of countries importing ornamental species rose from 32 to 120, with a slight dip in the interim. The largest of the importers of ornamental fish was Europe with 51 percent (the UK alone imports 19 percent of this figure), and North America with 26 percent of the market share (the USA making up 87 percent of this, making the largest single country importer with nearly 23 percent of the global market share).
Of the exporting countries, the fastest growth was seen in Czech Republic and Spain, while drops were noted in exports from the USA, Germany and Hong Kong, presumably linked to reducing imports into Japan, an important destination for ornamentals from these countries.


Monitoring of the global trade
The effective monitoring of the global trade is essential in order to properly record and analyse the volumes of species traded, in particular those perceived as vulnerable or under threat, to prevent irreversible damage. Through monitoring, a balance can be achieved and maintained between the demand for ornamental species, the need for income and employment, and the ecological requirements of habitats and populations. This balance can then provide for a sustainable industry into the future, reducing the risk of catastrophic loss of habitat or ecological imbalance potentially leading to socioeconomic issues in less developed areas.
In 2000, in response to a need for better monitoring of marine ornamental trade, the United Nations Environment Programme World Conservation Monitoring Centre (UNEP-WCMC), the Marine Aquarium Council (MAC) and members of various aquarium trade associations began, in collaboration, to address this need and created the Global Marine Aquarium Database (GMAD).
Trade data has been obtained from wholesale exporters and importers of marine aquarium organisms and integrated into quantitative, species-specific information which has been made public.


Fifty-eight companies, approximately one-fifth of the wholesalers in business, and four government management authorities have provided data to GMAD. In August 2003 the dataset contained 102,928 trade records (7.7 million imported and 9.4 million exported animals) covering a total of 2,393 species of fish, corals and invertebrates and spanning the years 1988 to 2003. It was believed that this data permitted the most accurate quantitative estimates available of the size of the global trade in marine ornamental fish and corals, and the first ever estimates for invertebrates other than corals.
A consultation on the monitoring the industry conducted in 2008 carried out for the European Commission by UNEP and the WCMC stated that a properly monitored and sustainably managed industry can present a valuable opportunity for income generation and support to livelihoods, while also providing an alternative to environmentally destructive activities.
Not monitoring the trade could, on the other hand, lead to an over exploitation of resources, damaging the long term future potential of the industry. The consultation identified the six mechanisms for monitoring the trade at species level as:

  1. the monitoring activities put in place by certification schemes (e.g. Marine Aquarium Council – MAC)
  2. GMAD
  3. the statistics generated by Customs and FAO,
  4. CITES
  5. veterinary controls
  6. Annex D of the EU Wildlife Trade Regulations.

In analysing these monitoring options, they determined that certification schemes are desirable but provide only partial coverage, are expensive, some have been unsuccessful, and there is little evidence of consumer awareness.
GMAD, being voluntary, was found to not be comprehensive enough for monitoring trade for conservation purposes.
Information generated by customs and FAO lacks the detail in the information required for conservation purposes.
CITES is effective at targeted monitoring of individual species of interest, however the monetary cost of obtaining permits to trade can be prohibitive.
Veterinary controls, for example in the EU, record species level data which could be useful for conservation purposes; however, at the time of the report, this data was being not captured and so valuable information was not being aggregated in a standardised and accessible manner.
Finally, Annex D of the EU Wildlife Trade Regulations proved to be the most effective tool for monitoring for conservation purposes, providing species level data of unrestricted species, with no monetary cost to the importer, making it the only instrument that could, at the time of the report, provide comprehensive species level data on the international trade in species of conservation concern. However, there was a willingness for EC veterinary controls to be investigated as a further mechanism for monitoring the trade.
While concerns were raised regarding the fact that these controls will only accurately monitoring imports into the EU while global trade may be underestimated, the fact that there is a system in place which is effective at monitor the ornamental trade is encouraging. It is therefore imperative that monitoring systems which can act on a global scale and based on those identified as being effective are initiated in order to provide proper traceability and sustainable development of the industry going forward.


Sustainability of marine ornamental supply
Despite sometimes being accused of causing undue degradation of populations and habitats, the marine ornamental trade is a low volume, high value industry. In 2000, 1kg of aquarium fish from the Maldives was valued at almost US$500, whereas 1kg of reef fish harvested for food was worth only US$6. Furthermore, the live coral trade is estimated to be worth about US$7,000 per tonne, whereas the use of harvested coral for the production of limestone yields only about US$60 per tonne (Wabnitz et al, 2003). There is therefore a clear financial incentive to preserve the important marine habitats and populations which provide to the ornamental industry, such as coral reefs and mangroves.
It is clear from the information available that the potential is there for a sustainable and profitable industry, but from the case study of the GMAD, there are clearly still large gaps in the knowledge on, in particular, marine ornamental harvesting.
This creates a need for a two pronged approach to developing a sustainable marine ornamental industry – the first being improved monitoring as discussed previously, and the second being an effort to increase the number of species which are cultured for the industry. Only one-10 percent of marine ornamental fish and less than one percent of hard corals are cultured (Wabnitz et al, 2003), this is in contrast to over 90 percent of freshwater ornamental species. In order to increase the proportion of marine species cultured, simple and cost effective culture methods must be sought which enable poor communities which rely on harvesting to switch their efforts to culture, thereby moving towards a more sustainable industry while not neglecting the beneficial potential of the ornamental trade for these communities.
The responsible aquarist
An appreciation by the home aquarist is the first step to self-regulation in terms of promoting sustainably sourced or farmed animals over those known to come from unsustainable wild fisheries. For example, in some wild fisheries collectors may use highly toxic substances such as sodium cyanide in marine environments and rotenone in freshwater systems to incapacitate the fish prior to collection.
Such practices can have long term toxic effects on the species assemblage and the community as a whole. Losses post capture can also be very high, up to 80 percent for some tropical marine fish, while other species such as cardinal tetra can have mortality as low as six percent, and so proper species selection to reduce demand for livestock which do not travel well can have a beneficial impact.
Additionally, better guidelines for collection, transport, and storage can help to reduce mortality. Therefore the consumer can have a marked impact on enhancing the sustainability of the industry through being aware of and choosing the most sustainably sourced livestock available, while ensuring they are properly educated on the requirements of their chosen livestock, so reducing mortality at home.
To highlight the role that responsible and properly informed aquarists can play, trade data, correlated with aquarium suitability information, indicates that two species known not to acclimatise well to aquarium conditions are nonetheless very commonly traded. They are the bluestreak cleaner wrasse (Labroides dimidiatus: 87,000 individuals traded between 1997 and 2002) and the mandarin fish (Synchiropus splendidus: 11,000 live individuals exported to the EU in the same period).
Data further indicates that species characterised as ‘truly unsuitable’, mainly due to their restricted dietary requirements, such as the foureye butterflyfish (Chaetodon capistratus), the harlequin filefish (Oxymonacanthus longisrostris) and the Hawaiian cleaner wrasse (Labroides phtirophagus), are also commonly traded, albeit in lower numbers (Wabnitz et al, 2003). Demand for species such as these is presumably perpetuated by mortality in home aquaria due to the unsuitable conditions, and it is these kinds of practises which can be minimised or eradicated through responsible aquarium keeping.
The global ornamental trade is a strong and growing industry, and it benefits all walks of life through wealth generation and aesthetic enjoyment. It has the opportunity to become a unique example of an ecologically and financially sustainable and renewable industry, where wealth flows from some of the worlds richest economies to some of the very poorest communities around the world.
However, in order to do this, improved systems for monitoring the global trade must be sought and implemented, and aquarists must strive to be as well educated as possible on the source and care of their livestock. In this way, the inhabitants of our home aquaria can remain some of the world’s most popular companion animals, while remaining affordable and healthy, and above all without damaging their natural habitats and populations.    ■

References
Livengood, E. J., & Chapman, F. A. (2008). The Ornamental Fish Trade: An Introduction with Perspectives for Responsible Aquarium Fish Ownership. University of Florida IFAS Extension, (FA124). Fisheries and Aquatic Sciences.
Ploeg, A. (2004). The Volume of the Ornamental Fish Trade. Ornamental Fish International. Ornamental Fish International.
Wabnitz, C., Taylor, M., Green, E. P., & Razak, T. (2003). From Ocean to Aquarium: the global trade in marine ornamental species. Cambridge: UNEP-WCMC.
UNEP-WCMC. (2008). Monitoring of International Trade in Ornamental Fish – Consultation Paper. Context.

EXPERT TOPIC – TROUT

Welcome to Expert Topic, a new feature for International Aquafeed. Each issue will take an in-depth look at a particular species and how its feed is managed. To kick off the first Expert Topic, trout takes centre stage. Over the next pages you’ll find, amongst other things, a feature on the trout value chain in Peru, a glimpse behind the scenes at Bibury Trout Farm in the UK and an overview trout culture and feed in Turkey.  First of all, industry experts from around the world give the inside track feed and management in their country. Enjoy.

Peru
by Anna Pyc, Peruvian Aquaculture Company, Peru
The highest industrial aquaculture center in the world is located in Peru’s central Huancavelica department, 4,600 meters above sea level. Peruvian Aquaculture Company (PACSAC) was founded in 2007 for the development of industrial aquaculture, an activity that is emerging worldwide as the main protein source for the near future.
To meet this goal, PACSAC integrates social and environment care with the use of modern technologies that make possible to provide quality products to international markets. At this stage, PACSAC is raising rainbow trout. As well as beign the higest trout farm in Peru, the company’s facilities are also the largest industrial aquaculture site in the country.
According to an article published by FIS.com, the company uses the same technologies applied in fish farming by the major producers of salmon and trout, like Norway, Chile and the United Kingdom.
However, this technology has been adapted to its unique environment and an individual model has been developed for the high Andes. The natural environment and the purity of the water in this mountain range is the greatest asset of the company, which allows making aquaculture a sustainable activity.
All procedures used by the company are environmentally friendly, and as a result, the Peruvian Aquaculture Company implements norms ISO 14001: 2004. PACSAC also develops regular environmental water monitoring to asses quality and sediment.The fish are fed exclusively with extruded fishmeal specially formulated for trout. After reaching the required market size the trout are harvested and transported to the processing plant.

Turkey
by Prof Dr Belgin Hossu, Faculty of Fisheries, Ege University, Turkey
According to the latest parametres from the Turkish Statistics Office, rainbow trout production is around 85.244 tonnes in Turkey and is increasing each day. Farms are mostly on land and damed lakes with a small amount in sea cages.
Rainbow trout production in Turkey can be divided in to two parts. The reason for this is the continously changing cost of feed due to fishmeal and fish oil prices. Unstable prices of fishmeal and fish oil has forced the farms to produce their feed themselves. As a result, the feed sector has to be seen as commercial feed mills and self-feed producers. Feed production for rainbow trout is done totally in Turkey. Foreign feed mills have built fish feed plants in Turkey because of increasing demand by farms. Trout farms choose feed mills according to their prices and payment ease.
Rainbow trout have differences between them because of geographic conditions of the country. Depending on these changing conditions, FCR is between 0.84- 0.97. The main protein source of feed is fishmeal. Fishmeal sources are foreign countries and sardine and anchovy from Black Sea according to the fishing seasons. Protein rate of feed is around 44 percent at growing ages. In addition to fishmeal, vegetable protein sources have been used to decrease the cost of the feed. These are soybean meal, corn meal and wheat meals. Adding of caroteneids generally happens at salmon breeding or matured fish of trout.
The feed manufacturing system is extruder. To decrease waste, floating feed is prefered.
Generally, the colour of the meat isn’t important because trout is consumed both fresh and frozen. On the other hand, yellow coloured meat isn’t preferable because it indictaes high levels of corn and soybean meal.

Poland
by Anna Py, Aller Aqua, Poland
Trout farming in Poland is situated mainly in the northern part of the country with its main species – rainbow trout. It is relatively young part of the Polish aquaculture reaching 14 thousand tons of annual production in 2010 with a value of approximately €40m. The farms are modern, many using partially recirculated systems and technology reducing environmental impact. Over 200 trout farms employ approximately 1000 people. The location of farms in rural areas makes them important for local employment levels.
Trout farms in Poland use high quality feeds purchased from leading feed producers in Europe. 41.5 percent of the market share belongs to Aller Aqua (2010), in the second place is Biomar (34.6 percent) and third is Skretting (12.3 percent). Aller Aqua is the only fish feed company with a production plant in Poland, which makes the company competitive regarding delivery conditions.
Trout farms in Poland are in most cases well managed and therefore the approximate FCR reaches values of starter feeds at 0.72 and, in fingerlings production 0.88. In recent years there has also been investment in automatic feeding systems to improve feeding effectiveness.
Very strict regulations in Polish law regarding the environmental impact of salmonids production also require that feeds that meet certain standards. According to individual farm water conditions the feed is chosen in respect of its caloric value and other properties. Many of the farms have their own hatcheries, where starter feeds are used. These feeds are especially important for having high survival rate and fish in good condition as a basis for fast growth.
Fish feed is the largest cost component of trout farms, amounting on average to about 34 percent, however it used to reach over 40 percent in previous years. Cost of labour, live raw material and other operational costs amount to 19 percent , 16 percent and 14 percent respectively. Other components costs are relatively small.
Polish trout production faces new challenges in terms of market demands as well as increasing pressure on reducing environmental impact. The Polish Trout Breeders Association faces these challenges and among other activities introduced a four year promotional campaign of trout in Poland. The campaign focuses mainly on promoting trout as a source of healthy nutrition, as well as spreading knowledge about the species.

Denmark
by Brian Thomsen, Director, Danish Aquaculture Organisation
Danish trout farmers almost exclusively use fish feed manufactured in Denmark. The mains reasons are that they view it as being superior and that it is in compliance with national legislation. Danish national legislation also regulates digestibility but most types of feed exceeds the legal requirements.
Key decision parameters when choosing a feed include: low FQR, high growth rates, national regulation, environmental impact – in general ‘price/performance’.
The national average FQR is approximately 0.94, thus we use on average approximately 940g of feed to make 1kg of trout. The protein content is gradually declining but it is typically around 42 percent. The main protein source is fishmeal.
The legislation for freshwater farming was changed this February. One of the key changes is that farms may now choose to be regulated on output (discharge quotas) and not on input (feed quotas). This will probably puts even more emphasis on the ‘price/performance’ ratio.
The fish farmers are well aware of the fact ‘that we are what we eat’. Therefore quality is of the highest importance. We mainly farm white fish but caratenoids are used in fish feeds that are used to make pink trout.
The cost of feed is always a key factor but the cost is judged against performance. The key question is therefore not the price per kg per se but the ‘price/performance’ ratio.
We produce approximately 10,000 tons of trout in marine farming and 25,000 tons in freshwater farming. The gross output (2010) was approximately €45m for marine farming and €80m for freshwater farming. The industry employs approximately 1000 people (including production and feed and processing).

United Kingdom
by David Bassett, British Trout Association
UK trout farming differs to some other countries in that the UK employs a number of different production methods. Trout are farmed in freshwater open net pens, earth ponds and concrete raceways and are also farmed in open net pens in marine water off the west coast of Scotland. UK trout farmers also employ recirculation technology – most commonly as partial recirculation in hatchery facilities rather than the entirely closed recirculation sites as may be seen elsewhere.
The UK primarily produces rainbow trout, although brown trout are farmed too. Both species may be farmed to organic standards, and consequently use organic feeds, although this market remains small, producing only in the hundreds of tonnes. Both brown and rainbow trout are farmed for the restocking market (i.e. sale of live fish for stocking to fisheries) although the majority of fish that are farmed are for the table market.
Production tonnages vary annually, but current official statistics suggest that circa 11,000 tonnes of table trout are farmed each year, with a further circa 3,500 tonnes for restocking. Large trout production, those fish farmed in marine water, is increasing, with 2011 production being estimated at 2,000 tonnes, up from circa 1,600 tonnes in 2010.
Fish feed accounts for approximately 50 percent of production costs, and so is of paramount importance to UK producers. Through both European Union and UK domestic legislation, fish feeds, their composition and their use, are tightly regulated. The vast majority of trout farms source feed from the major commercial suppliers. Skretting has the largest market share, although other suppliers include EWOS, Biomar, Le Gouessant and Aller Aqua. However, whilst costs are high, trout farmers seek value for money and a return in terms of performance and as such would prefer to pay for a top quality feed in that this is a better investment in the long term resulting in a better yield and healthier fish.
Feed compositions vary between manufactures and specific formulations / diets. The major source of protein continues to be fish meal. Increasingly, producers seek to be able to vary the inclusion rates in diets of such ingredients as fish and vegetable oils. With the global commodity index affecting the price of key ingredients, trout farmers support feed manufactures in their attempts to operate using as wide a basket of ingredients as possible, to optimise variations in the commodity market.
With the exception of fish farmed to organic standards, the UK market prefers fish that is “pink” fleshed. As such, astaxanthin on canthaxanthin are included in the formulation of diets.
Most UK feeds for the table market avoid using land animal protein. Although permitted to do so by law, retail buyers seem reluctant to purchase fish fed using such diets. However, research undertaken by industry and other third parties suggests that there is little to no opposition to the inclusion of such protein sources on the part of the consumer / general public, who remain generally unconcerned about the diets fed to farmed fish.
In common with other sectors, ‘sustainability’ is a term that is used increasingly often with regard to fish farming and fish feeds. Whilst a definition of sustainability is always hard to achieve, it would be fair to suggest that much greater emphasis is now being placed upon such issues as Fish In Fish Out (FIFO). As a trade association representing the UK farming industry, the British Trout Association is increasingly liaising with feed companies and NGO organisations over issues relating to the inclusion percentages of fish meal and fish oil in diets, and the origin of the fish meal and fish oil that is used. It is predicted that greater emphasis will be placed upon such issues in the future, with certain certification schemes placing greater emphasis on the sustainability imprint of all aspects of production.  How much importance consumers attach to this has yet to be demonstrated.
UK fish farming is strictly regulated in relation to discharges into the aquatic environment. As such, farmers pay close attention to feed conversions ratios and associated nutrient discharge and suspended solids. Whilst feed conversion ratios vary across the UK, given the wide range in production systems, water temperatures and other variables, feed conversion and feeding protocols have continued to improve in sophistication and understanding with reported ratios varying from under 1:1 (typically 0.95) to 1.2 :1.
UK trout farmers enjoy a close and mutually beneficial working relationship with commercial fish feed manufacturers and as an industry we continue to work together to be at the forefront of trout production.

Buxton Trout and Salmon Farm, Australia
by Mitch MacRae, Secretary of the Australian Trout & Salmon Farmers Association
At Buxton Trout & Salmon Farm we have a general feed conversion rate of 1.1 – 1.2. Protein content of the feed we use is 45 per cent and the percentage of fish meal is not known. There are only 2 feed suppliers in Australia, we chose Skretting as their feed performs better (better FCR) without compromising environmental targets on discharge and is more economical because of freight costs.
The taste of the trout is not affected by the feed, however the colour is depending on how much colour is added to the feed. Costs of feed and FCR are important factors as margins are tight, and fish feed is one of our biggest costs. On average to produce 1kg of trout we will need 1.1 – 1.2kg of feed.
In Australia approximately 1500-2000 tonnes of trout are produced each year, with approximately 85 per cent of Australia’s trout being grown in the Murrindindi region in Victoria. Australia’s trout production has an approximate value of 10-15 million dollars per year and employs approximately 200 people.
Trout culture and feed in Turkey
by Dr Atilla Ozdemir, Central Fisheries Research Institute, Turkey
Although Aquaculture has a relatively short history in Turkey, it began with rainbow trout (Onchorhynchus mykiss) and common carp (Cyprinus carpio) in the late 1960s and developed further with gilthead seabream (Sparus aurata) and European seabass (Dicentrarchus labrax) culture in the mid-1980s. Production reached 167,000 tonnes a year in 2010 of rainbow trout, seabass, seabream, mussel, common carp and other species, produced on nearly 2100 farms.
The rainbow trout has been cultured since the early 1970s and Turkey has become one of the top trout producing countries in Europe with an annual production of over 85,000 tonnes, or almost 50 percent of the country’s total aquaculture production. With the surprising appropriate ecological supply for trout culture in the marine environment thanks to low salinity the Black Sea has an enormous potential. Today there are more than 20 sea-based farms which are situated in the Black Sea. This tends to increase in number of fish farms and production.
Apart from marine and some freshwater cage farms in lakes and reservoirs, the majority of the trout farms employ small concrete raceways mainly using stream waters. In the past ten years, trout cage culture in dams has reached a very important level of production. Over 50 percent of the farms have their own hatcheries with eggs being produced during the natural breeding season (between December and February). Ongrowing in raceways lasts between 12 and 24 months. The majority of fish are sold locally as portion size white trout.  In the Black Sea, fish are reared in cages up to 0.5–1.5 kg and sold as ‘Black Sea salmon’.
Steadily increasing production has accompanied a large volume of fish feed needs. Trout feeds are produced in state-of-the-art facilities using leading-edge quality assurance techniques. There are currently 10 feed mills with the total capacity of over 300,000 tonnes per year. Almost all feed mills produce trout, sea bass and sea bream feeds using extruding technology made after 2000. Since the regulatory standards are high, all fish mills track  raw materials acquisition, handling and storage, production processes and packaging and delivery.
The main protein source is always declared as fish meal. But reliable data on this is hard to obtain. Although Turkey has different zones all around country having various water characteristic features, the feeds are produced regardless ecological differences as implemented in some countries by chosen different protein/lipid/energy ratios.
There is high variation in FCR depending on feed management in farms and location of farms. The lowest rate obtained in cages in Black sea as 0,9 and highest as 1,2 in inland farms. The effect of type of feed on taste and colour of the fish is has not been considered very much so far.
The initial on-farm experience and following demands/complaints from processing units may lead producers to select the best available feed. No carotenoids are incorporated into most trout diets produced in Turkey but in some cases producers in Black Sea demanded pigmented feeds.
The cost and quality are almost equal factors in choosing feed. As production increases, the market competition is getting more stressful for producers. As a consequence of competition between farmers, feed producers are always introduced the best available feeds in order to reach desired size as quickly as possible. So the growth rate is almost primary factor feed driven. The intrabrand competition occurs also among feed producers.
The environmental pressures and impacts caused by the typical production of rainbow trout in Turkey have been taken into consideration particularly in the last decade. After the adoption of new regulations on aquaculture in Environment Law, all aquaculture facilities are under a monitoring programme. Through implementation of a control programme the farmers are directly or indirectly forced to use better quality feeds particularly in low phosporus content. Since the overall production is increasing steadily, this pressure is expected to increase in near future.

Behind the scenes at Bibury Trout Farm – A working trout farm that is attracting new business from tourism
by Kate Marriott, General Manager, Bibury Trout Farm, United Kingdom

Bibury Trout Farm is one of Britain’s oldest, and certainly most attractive, trout farms. Founded in 1902 by the famous naturalist Arthur Severn, the farm was set up to stock the local rivers and streams with the native brown trout. Today, it covers 15 acres in one of the most beautiful valleys in the Cotswolds, the Coln Valley. The Farm has diversified over the years, and the leisure side of the business now plays a very important part.
Situated in the heart of the beautiful village of Bibury, the farm benefits from the large number of tourists who visit the region.The crystal clear waters of the Bibury Spring provide the essential pure water required to run the hatchery which spawns up to six million trout ova every year.
Up to a third of the ova are sold to outlets throughout Britain and occasionally abroad.  The remainder are grown on and sold to supply angling waters throughout the country, approximately 80 tons.  A small proportion (20 tons) are sold to other trout farms to supply the table market and are sold to local hotels, wholesalers and to the public through both the farm shop and farmers markets as fresh gutted trout, fillets or smoked trout, all smoked and packaged on the premises.
On the farm visitors can learn about the rainbow and brown trout while they wander in the beautiful surroundings. There is a chance to see grading in progress when the fish are selected for size and quality before being transported to new homes in oxygenated water in specially made fibre glass tanks.
Information boards give a insight to what goes on in the hatchery and fryary areas and staff are on hand to answer any questions. Feeding is done daily by staff and the water comes to life as the fish vie for the last morsel.
For the more adventurous, or the budding fisherman, Bibury’s Catch Your Own Fishery is an ideal opportunity to catch your supper or get hooked on a new hobby. Open at weekends during March – October, and during the local school holidays, we provide all the equipment and help if required.
In addition to the farm, our recently refurbished fish shop which now houses a wonderful range of wines, deli products, and preserves as well as quality breads, eggs, and milk.
The Trout Farm is situated in the centre of the village of Bibury, next to Arlington Mill. Bibury is between Cirencester and Burford in the United Kingdom
Website: http://www.biburytroutfarm.co.uk
Emerging disease in Mexican trout
by Celene Salgado Miranda, Mexico State University, Mexico
Infectious pancreatic necrosis (IPN) is a disease caused by a birnavirus affecting several wild and commercial aquatic organisms. Salmonid species are the most affected, having an important impact in the salmon and trout culture due to a high rate mortality of fry and fingerling. IPN disease is listed in the fish diseases of the International Health Code, World Organization for Animal Health (OIE). For this reason, any IPN outbreak has to be reported.The epizootiological knowledge of the IPN is relevant for establishing preventive and control strategies against both disease and causative agent.
Distribution
The IPN and the causative agent (IPNV) has been reported in several countries: Australia, Canada, Chile, Denmark, Scotland, Spain, Finland, France, England, Italy, Japan, Norway and Switzerland, among others. Based on these reports, IPN is regarded as a worldwide distributed disease. In Mexico, IPNV was identified in 2001 from US-imported rainbow trout fry. In a recent study, the IPNV was isolated from three rainbow trout breeding farms located at Mexico State, Mexico, regarded as the main producer of this fish species.

Etiology
The causative agent of IPN is a virus belonging to the Birnaviridae family. Other members of this family include infectious bursal disease (IBD) of chickens and X virus of Drosophila melanogaster. This birnavirus is single-shelled icosahedrons with characteristic isometric hexagonal profi les and has a diameter of about 60nm. The genome consists of two segment of double-stranded RNA. Genome segment A encoding two structural proteins (VP2 y VP3) and a nonstructural protease, while segment B encoding for a RNA polymerase. VP2 protein induces the production of specific-type neutralizing monoclonal antibodies. It is thought that VP2 contains all the epitopes recognized by these antibodies.
The serological classification scheme of Hill and Way recognizes nine different IPNV serovars into the serogroup A. Seven of these serotypes have been identified in IPNV rainbow trout isolates. Serogroup B includes a single serotype represented by the TV-1 archetype isolated from brown trout (Salmo trutta) and common carp (Cyprinus carpio). Each serotype includes a number of strains that differ in virulence. This variation complicates the disease which is little understood.

Epizootiology
Natural and experimental hosts Salmonids are the most susceptible species under natural conditions. The brook trout (Salvelinus fontinalis) is the most susceptible one to lethal effects of IPNV, followed by rainbow trout (Oncorhynchus mykiss) and Atlantic salmon (Salmo salar). Also, IPNV has been isolated from artic char (Salvelinus alpinus),brown trout (Salmo trutta) and lake trout (Salvelinus namaycush).
The IPNV has been isolated from important non-salmonid species in marine aquaculture: turbot (Scophthalmus maximus), sole (Solea senegalensis) and Atlantic halibut (Hippoglossus hippoglossus).Also has been isolated from some fishs as pike (Esox lucius), goldfish (Carasius auratus), discus fish (Symphysodon discus) and bream (Abramis brama), among others. IPNV was experimentally inoculated and re-isolated from zebra fish eggs (Brachydanio rerio).Some IPNV cases have been reported in American and European eels (Anguilla anguilla). However, the infection in Japanese eel (Anguilla japonica) has a greater economic impact.
In summary, the IPNV has been identified in a number of teleosts family fish: Anguillidae, Atherinidae, Carangidae, Channidae, Cichlidae, Clupeidae, Cobitidae, Cyprinidae, Gadidae, Esocidae, Percichthyidae, Percidae, Pleuronectidae, Poeciliidae, Salmonidae, and Sciaenidae.
Transmission, carriers and vectors
Infected fish can transmit the virus by both horizontal and vertical transmission. These fish shed the virus by urine and faeces, contributing to the horizontal transmission. In breeding fish, it has been demonstrated that the IPNV is vertically transmitted by viral adsorption to the surface of spermatozoids, or it can be present in the follicular fluid, but not in the nonfertilized eggs.
Bebak et al. experimentally determined the IPNV excretion patterns in rainbow trout fry. The time between challenge and excreting, and challenge and signs onset were evaluated.
The authors also estimated the rate of susceptible-excreting fish into a population from inoculated IPNV fry. It was demonstrated that IPNV-infected rainbow trout fry shed the virus two days post-inoculation, and the shedding is increased, and approximately decreased after 12 days post-inoculation. More than 75 percent of the rainbow trout population was infected in less than a week from the beginning of the viral shedding.
In rotifers (Brachionus plicatilis) it has been observed birnavirus lesions associated with an IPNV-like virus. It is likely that invertebrate animals used as living-food for seabream and turbot larvae, could be involved in the viral transmission. Similarly, it has been demonstrated that the freshwater crayfish (Astacus astacus) retains the virus in tissues and hemolymph, constantly shedding the virus to the water. Halder and Ahne suggest that these organisms are infected by the consumption of IPNV-infected trouts.
The following shellfish species are regarded as reservoirs of the IPNV: mussels (Mytilus galloprovincialis), oysters (Crassostrea gigas), periwinkles (Littorina littorea), and wild fish as sand eels (Ammodytes sp), sprat (Sprattus sprattus) and blue whiting (Micromesistius poutasou), among others. IPNV has been also isolated from moist fish pellets and marine sediments. Wild piscivorous birds are regarded as vectors of the IPNV, which can be isolated from faeces samples.
Signs
The IPN is a typical disease in early ages of salmonids, causing up to 100 percent of mortalit y in fi ngerlings and fi rst-feeding fry. An experimental study reported a mean cumulative mortality ranging from 84 percent to 92 percent in challenged Atlantic salmon fry. The fish mortality started seven days post-challenge and peaked at 10-12 days.
Generally affected fish showed anorexia and rotate about their long axis in a whirling motion with lapses of ataxia. In these fish darkening occurs (hyperpigmentation). Mild to moderate exophthalmia and abdominal distention are common. Also, gills are typically pale and hemorrhages are sometimes present in ventral areas, including the ventral fins. Many emaciated fish trail long, thin, whitish, cast-like excretions from the vent.
Macroscopic and microscopic findings
According to necropsy findings, spleen, heart, liver and kidneys of fry are abnormally pale and the digestive tract is almost always devoid of food. Petechiae are observed in some viscera. Sometimes, food residue remains in the gut, the quantity is small and confined to the far distal or rectal portion. Very often the body cavity may contain ascitic fluid. The stomach and anterior intestine contains a clear to milky cohesive mucus, among other findings.
Main lesions found at the histopathology study include: focal coagulative necrosis in pancreas, kidney and intestine. The pancreatic tissue showed degenerative changes, including acinar cell areas, and zymogen granules freeing. Nuclear pyknosis of different sizes are observed. In many cases, inflamatory cell infiltration is not evident. In fish that suffered the disease up to two years before the histology study, hypertrophy of Langerhans’ islets with abundant fibrosis were found.
In cases of pancreatic lesions, also acute enteritis featured by necrosis and sloughing of the epithelium are observed. In the intestinal lumen, catarrhal whitish exudate is associated with the disease. Inclusion bodies are not observed in affected cells. In many cases, the renal tissue has small focal degenerative changes. In fish that were infected during early ages, abundant rounding up of epithelial cells with karyorhectic nuclei was found. This finding suggest that they can be viral replication sites in carrier fish; however, it has not been confimed.
Diagnosis
The procedure for IPN diagnosis, recommended by the OIE, is based on the isolation of IPNV in susceptible cell lines (Figure 1), and further identification by serological techniques by immunofluorescent test, neutralization test and ELISA.
Diagnosis of clinical outbreaks is based on histology and immunological ev idence of the IPNV in infected tissues. These cases are confirmed by the IPNV isolation and immunological identification of the virus. Due to insufficient knowledge of the serological responses of fish to IPNV infection, the detection of fish antibodies to IPNV has not been accepted by the OIE (2003) as routine tests.
Detection of IPNV in cell lines is consistent and simple, particularly in cell lines from homologous species. It is due to: 1) the virus is present in high level titers in the tissues; 2) viral isolation could be positive from non-diseased fish 3) viral isolation could be positive from any viral phase; 4) two to three weeks are required for isolation and identification of the agent, which is not a critical issue for presentation of a epizootic outbreak, and 5) high sensitivity and easy observable cytopathic effect. Cell lines used for the IPNV isolation include: RTG-2 (rainbow trout gonad), CHSE-214 (chinook salmon embryo) and BF-2 (bluegill fry).
Currently, some methods have been developed for detecting IPNV by reverse transcriptase-polymerase chain reaction (RT-PCR) technique. However, sensitivity of this technique has not been greater than the cell culture. Hence, viral isolation and serological confirmation of the virus are regarded as the choice procedures for the IPNV identification.

Prevention and control
Current preventive methods are based on the onset of control and hygiene practices during rearing of salmonids, avoiding introduction or importation of fertilized eggs or fish from IPNV-infected breeding trouts. Also, the use of fish-free freshwater (for example, spring water), particularly IPNV-carrier fish, reduces the risk of infection. However, in Mexican trout farms, this condition is not always possible. As mentioned above, Salgado-Miranda carried out the IPNV isolation from three rainbow trout breeding farms located at Mexico State.
Obtained results indicated a possible horizontal transmission throughout the water supply from a farm where a previous IPN outbreak in fry was recorded. In these cases, treatment of supplied water could decrease the risk of an IPN outbreak and other infectious agents. Liltved et al. experimentally exposed live cultures of Aeromonas salmonicida subsp. salmonicida, Vibrio anguillarum, V. salmonicida, Yersinia ruckeri and IPNV to ozone or ultraviolet (UV) irradiation at nine°-12°C. The four bacteria tested were inactivated by 99.99 percent (fourlog reductions in viable count) within 180 seconds at residual ozone concentrations of 0.15-0.20 mg/L.
The IPNV was inactivated within 60 seconds at residual ozone concentrations of 0.10 a 0.20 mg/L.
Similarly, the four bacteria tested were inactivated by 99.9 percent (five log reductions in viable count) at a UV dose of 2.7 m Ws/cm2 at room temperature. IPNV was much more resistant to UV irradiation than the bacteria. An average UV dose of 122 m Ws/cm2 was required for 99.9 percent (three log) reduction in virus titer. However, it has to be considered that ozone low residual levels (0.010 a 0.20 mg/L) have also caused mortalities in trout recirculating systems.
A concentration of 40ppm available chlorine was required to experimentally inactivate 10 TCID50 of IPN V/ml in 30 minutes. Similarly, a concentration of 35ppm of active iodine was required to completely inactivate 10 TCID50 of IPNV/ml in the same time. Other studies, where several disinfectants were tested, 25ppm of iodine was required to inactivate IPNV, infectious haematopoyetic necrosis virus (IHNV) and viral haemorrhagic septicemia (VHS).
It is important to highlight that IHNV and VHS are exotic infectious agents in Mexico. For controlling IPN in breeding farms, infected fish and its offspring (eggs, fingerling and fry) have to be sacrificed. IPNV transmission by fertilized eggs can occur in spite of iodine treatment. Propagation of IPNV-free stocks monitored by viral isolation during several years, has been a good strategy for the control of IPN in breeding farms.
In areas where IPN is enzootic, it is recommended, during an outbreak, to decrease the density of the affected population, reducing the impact on the total mortality. A study showed that interaction between fish density and number of infected fi sh, affected signifi cantly the mortality parameter. However, there are some disagreements about it.
Up to date, highly effective IPNV-inactivated vaccines do not exist. Treatment with formalin or ß-propiolactone for use in vaccines, completely inactivated IPNV, but caused a slight reduction in antigenicity up to 50 percent . An active vaccine containing an IPNV non-pathogenic strain, normal trout serum-sensitive, did not confer protection in experimental challenged fish.
In Norway, both inactivated and recombinant vaccines are widely used. The recombinant vaccine, the first one licensed for using in fish, express the VP2 sequence in Escherichia coli and induce specific IPNV antibodies.
As it happens in other viral diseases, there is no treatment for the IPN. Several antiviral compounds inhibits the in vitro replication in cell culture; for example, ribavirin, pyrazofurin and 5-ethynyl-1-ß-D-ribofuranosylimidazole-4-carboxamide (EICAR),among other compounds. Research on EICAR as an antiviral compound showed good results in experimentally IPNV-infected rainbow trout.
The effect of the administration of lysozyme (KLP-602) in the feed of IPNV experimentally infected rainbow trout, has been also evaluated. Cumulative mortality was lower in fish fed on dietary treatment containing lysozyme (30%), compared with untreated fish (65%). Based on the significant increase of all the immunological parameters, these authors refer that the lyzozyme modulated the cellular and humoral defense mechanisms after suppression induced by IPNV. Also, a selected trout strain resistant to natural infection by this virus has been reported.
As Håstein et al. pointed out, future national and international aquaculture regulations for the establishment of preventive and control strategies of infectious diseases include: adoption of standardized control methods, suitable infrastructures development, and a deeper comprehension of the epizootiology of aquatic organism diseases.

Conclusion
IPNV is a birnavirus affecting mainly salmonid species, being the rainbow trout the most susceptible species. In Mexico, isolation and identification of this infectious agent from rainbow trout was recently reported. Neither a treatment nor totally effective vaccines against this disease are available, being the preventive and control measures of great importance. Introduction into farms of eggs, fish and water supply free of IPNV are the main preventive strategy. These also constitute the most important risk factors in spreading of this disease.
Acknowledgments
The collaboration in structure design and critical review of the manuscript by Dr. Edgardo Soriano Vargas, CIESA-FMVZ-UAEM, is greatly acknowledged.

Struggling Downstream? – The trout value chain in Peru
by Jodie Keane with Alberto Lemma, based on studies by Juana Kuramoto at GRADE, a member of the Consorcio de Investigación Económica y Social (CIES), Peru.
In Peru, the United States Agency International Aid (USAID) project for Poverty Reduction and Alleviation (PRA) has been one of the pioneers of value chain interventions. Under the PRA, value chains of distinct products have been fostered, ranging from agro-industrial products to artisanal goods and small manufacturing, which have then gained placement in international markets.
The chain for trout is one of the successful chains achieved by the PRA. Not only has it combined the natural advantages of raising this fish in Peruvian territory, but also it has managed to consolidate access to foreign markets through a national producer and trader, Piscifactoría Los Andes.
Raising trout has a long history in Peru. The species was introduced in the country in the 1930s, with the import of eggs and fry brought from the US. The development of trout farming occurred extensively, by populating lakes and water sources.
By the 1980s, there was a new effort to propel this activity through the construction of fish farms in various mountainous provinces of the country. However, raising trout did not take off as an economic activity and the infrastructure that was constructed was left underused.
In the 2000s, the Peruvian enterprise Piscifactoría Los Andes made important efforts to begin trout export to foreign markets. These efforts complemented the PRA project, with the development of trout value chains initiated in Junín, Huancavelica and Puno.
Linking producers to exporters
Although the initial investments required for trout production may be low, export of trout to international markets requires a series of sanitary certifications, imposing a high cost on producers and traders. The value chain for trout is divided into three well-determined links: fry production, trout production and marketing. These links define the principal actors in the value chain.
One company in Peru accounts for the majority of trout exports (90 percent as of 2006) and is the largest and oldest within the industry in Peru. Piscifactoría Los Andes recognised that it would need to increase production in order to begin exporting trout. In 2000, the company decided to participate in the PRA project and initiated negotiations with producer organization, SAIS. The company would provide the necessary capital as well as purchasing the fry and balanced food.
SAIS agreed to hand over its production to the company once the trout had reached the optimal size and weight. The PRA financed the contracting of several experts, who provided technical assistance to SAIS.
Despite initial incompliance on the part of SAIS, the interaction allowed for Piscifactoría Los Andes to increase production and sell to the export market. The agreement between SAIS and Los Andes was broken, but the coordination model must have appealed to the company because it continued to participate with the PRA in other regions. In fact, the company has signed an agreement with private company California’s Garden de Oxapampa within the framework of a PRA project.
The experience of Acoria
In this case, the coordination was between the Municipality of Acoria and the Los Andes company. The agreement continues to present day. In 2003, initial production reached 12 metric tones of trout per year and production is expected to reach 72 metric tones in 2008. In 2007, the municipal enterprise attained financial self-sufficiency and managed to generate employment for its community members (including single mothers and widows). The Municipality of Acoria is contemplating initiating trout farming in other locations in its jurisdiction.
There are several lessons to be learnt from this experience. First, coordination agreements need to be put in place to facilitate investments in public infrastructure; management could be realised by a municipal company or by a company under a franchise agreement. Second, it is feasible to replicate coordination models on a small scale, but it is still necessary to include components of technical assistance and financing. Third, expanding the level of production of trout requires significant capital. The financial solvency of large producers and coordinators is vital.
The experience of Puno
In Puno, there are different institutions linked to trout farming, such as strong producer associations like the Association of Trout Producers (APT), which are relatively active in promoting technical assistance projects for the benefit of their associates. However, initiatives have not managed to coordinate the value chain strongly, owing to budget limitations and overemphasis on the provision of basic training to the neglect of other activities. The low prices that are prevalent in the region, lack of credit, high levels of informality, lack of coordination of the actions of state organs and scant knowledge of marketing aspects of trout are the main obstacles to the development of this activity in Puno.
Conclusions
This study showed that value chain interventions should be utilized for programs whose main objective is to increase the dynamism of economic activity in a specific territory, as such programs are not necessarily effective in alleviating poverty.
In general, value chain interventions are targeting foreign markets, which are subject to quality certification and sanitary norms that can present bottlenecks for small local producers. Moreover, coordinating the chain requires significant technical and financial capacity. In the examples discussed, such assistance has been forthcoming from the PRA, but project objectives have not always been achieved.
It is important not only to provide technical assistance, but also to offer access to finance and to facilitate institutional development. Sensitisation programs are recommended in order to promote the formalization of producers and membership in associations, and to engender confidence and respect in the agreements.
Poverty alleviation programs should be designed mainly to elevate basic poverty indicators, and not to coordinate with sophisticated markets. Poor producers generally manage a range of resources and activities in order to support themselves, and often consider focusing on a single economic activity to be a high risk.
In parallel with the promotion of value chains that coordinate with foreign markets, it is necessary to work on the formation of value chains that coordinate with regional markets and the domestic market, in order to prevent prices falling from excess supply. To this end, it is necessary to work on the formation of regional markets and the provision of public goods in the form of physical infrastructure and market information systems.
The focus on demand promoted by the PRA project should be supported by the important activity of market intelligence. Only in this way will we be able to construct stable demand for local producers and ensure that market prices are adequate in order to generate sufficient utility to cover the risk that they face for their specialization.

The origin of introduced rainbow trout – in the Santa Cruz River, Patagonia, Argentina, as inferred from mitochondrial DNA

by Carla M. Riva Rossi, Enrique P. Lessa, and Miguel A. Pascual. Centro Nacional Patagónico (CONICET), Uruguay

Salmon and trout have been transplanted to habitats throughout the world and self-sustaining populations have been successfully established globally, with the exception of Antarctica (MacCrimmon 1971; Quinn et al. 1996; Nielsen 1996). Rainbow trout (Oncorhynchus mykiss) was first introduced into Argentinean Patagonia, the southernmost region of South America, at the turn of the twentieth century and eventually became the most conspicuous freshwater species in major river basins of the region (Pascual et al. 2002b).
Like all other known introduced rainbow trout around the world, typical Patagonian fish remain in fresh water throughout their entire life cycle, with a life history similar to that of resident populations in rivers and head lakes in western North America (Wydosky and Whitney 1979). The Santa Cruz River in Patagonia (50°S) is the only drainage in the world where introduced rainbow trout are known to have developed partially migratory populations composed of individuals exhibiting a marine migratory phase, so-called steelhead, and strictly freshwater fish that remain resident in their native stream (Pascual et al. 2001).
As in many other salmonid populations with this dual anadromous–nonanadromous life history, the way and extent to which the two ecotypes intermingle in the Santa Cruz is uncertain. Genetic analyses based on microsatellite loci revealed that the anadromous form is genetically indistinguishable from main-stem resident trout (Pascual et al. 2001), suggesting that significant gene flow occurs between the two forms.
Whether the introduced fish were in effect anadromous or anadromy arose in situ remains unknown (Behnke 2002; Pascual et al. 2002a). We also ignore the specific mechanisms underlying the expression of alternative life histories in the Santa Cruz, i.e. a genetic polymorphism, a genetically determined developmental threshold (i.e. the link between individual growth performance and anadromy or nonanadromy; Thorpe et al. 1998), or an entirely environmental effect. At this point, there are critical aspects regarding the environmental versus genetics bases of life history variation in Santa Cruz River rainbow trout that we do not know.
A logical first step to start elucidating the bases of life history variation in Patagonian rainbow trout, in particular, the development of anadromy, is to assess their genetic legacy through the identification of the parental sources. Poor historical bookkeeping and complex ancestry have made it difficult to address this issue from transplant records alone.
The Santa Cruz River, as well as all other rivers throughout Patagonia, received rainbow trout from two main sources at different times. Between 1904 and 1910, rainbow trout ova were imported from the United States (US), most likely derived from rainbow trout and steelhead from locations in northern California or southern Oregon (Pascual et al. 2001,2002a; Behnke 2002). After the 1930s, and particularly after the 1950s when fish transplants within the region became more common, all rainbow trout plantings were based on new stocks imported from Germany and Denmark (Baigún and Quirós 1985). However, the Santa Cruz River has had a history largely independent from that of more northerly Patagonia locations, with only occasional introductions after 1920 (Pascual et al. 2001, 2002a). Thus, presumably, wild populations in this river were mostly derived from the early shipments from the United States.
Mitochondrial DNA (mtDNA) has proven very successful for identifying the origins of several introduced salmonid populations and for assessing genetic differences between contemporary wild and introduced populations (Quinn et al.1996; Burger et al. 2000). In this paper, we use mtDNA sequence variation to identify the founding populations of Santa Cruz River rainbow trout. We start by analysing mtDNA sequences of both resident and migratory fish. We include in the analysis fish from a local hatchery, which was founded with European trouts widely stocked around the region after 1950.
We then build and apply a probabilistic model of random survival and reproduction of individual fish to calculate the likelihood that wild Santa Cruz fish had originated from a collection of candidate North American stocks. Finally, we discuss the merits of the techniques applied to evaluate the relative contribution of pre-1950 transplants from US stocks and post-1950 transplants from Danish stocks to wild populations of rainbow trout throughout Patagonia.
Transplant history
From 1904 to 1910, several consignments of rainbow trout embryos arrived in Argentina, mainly from the United States, with only occasional imports from European countries, such as France and Germany (Tulian 1908; Marini and Mastrarrigo 1963; Behnke 2002). Between 1906 and 1910, a total of 105,000 rainbow trout ova collected in the United States were shipped to the Santa Cruz River. 25,000 in 1906, 30,000 in 1908, and 50,000 in 1909. The 1908 shipment was completely lost, but the other two consignments were successfully hatched and planted in the river, with comparable losses throughout (about 65 percent; Tulian 1908; Marini and Mastrarrigo 1963). For practical purposes, the number of eggs from the parental populations giving rise to the Santa Cruz stock was 75,000.
The most likely origin of these eggs was the Baird Station on the McCloud River, California (Pascual et al. 2001). However, they may as well have come from steelhead and rainbow trout in alternative northern California and southern Oregon locations (Behnke 2002; Pascual et al. 2002a). Rainbow trout introductions into Argentina intensified after 1950, this time based on stocks from Denmark (Pillay 1969; MacCrimmon 1971) and maintained by Bariloche.


Northern Patagonia hatchery
By that time, Bariloche became the main center of salmonid propagation in Argentinean waters, contributing to the distribution of these new stocks throughout the 1950s, 1960s, and 1970s. Danish stocks of rainbow trout have a complex ancestry; multiple lineages from California, Michigan, Canada, New Zealand, and France appear to have contributed to their foundation (MacCrimmon 1971). Small consignments of these fish(<2000 embryos) arrived at the Santa Cruz River from Bariloche in the 1970s and were planted in second- to-third order tributaries flowing into the upper basin (Fig. 1).


Finally, in 1991, the Piedra Buena Hatchery was built on the lower Santa Cruz River (Fig. 1). The fish used to found this hatchery’s broodstock also came from Danish fish, as those kept by the Bariloche Hatchery.
Although fish of this hatchery are not used for stocking the river, escapes are likely, so that some introgression with wild fish might occur (Pascual et al. 2001). In any case, these fish provide a representative group of known Danish origin with which to contrast the genetic structure of Santa Cruz
River wild fish
The upper Santa Cruz basin is dominated by two large glacial-fed lakes, Viedma and Argentino, that form the Santa Cruz River. The main stem river has an average flow of 690 m3·s–1 and extends for 382 km across the Patagonian plateau draining into the Atlantic Ocean (Fig. 1). Landlocked populations of rainbow trout inhabit most of the second- to third-order tributaries that feed the head lakes; few springs and small tributaries enter the main-stem river, none of them significant from the point of view of their trout populations.
We restricted our analysis to the main-stem river populations, which, as revealed by a telemetry study, is the domain of the anadromous rainbow trout and of the resident fish to whom they are most likely related (Riva Rossi et al. 2003). Adult anadromous and resident rainbow trout were caught by hook and line and by gill nets between 2000 and 2002 in April and September along the main-stem Santa Cruz River (Fig. 1).
Sampling locations were based on spawning and fishing abundances documented in previous surveys and consisted of two river reaches located in the upper course (‘Primer Laberinto’ and ‘Segundo Laberinto’), one in the middle course (‘Ea. Rincón Grande’), and one in the lower course (‘Cte. L. Piedra Buena’ City).
At each locality, tissue samples were obtained from fish of each ecotype. From a total of 182 wild fish captured, 20 were successfully sequenced: five individuals of each ecotype from the upper course, three resident fish from the middle course, and three anadromous and five resident fish from the lower course. Direct inspection of external characteristics and scale pattern analysis were used to distinguish anadromous from nonanadromous fish (Pascual et al. 2001). Also, fin clips were obtained from five spawners from the Piedra Buena Hatchery broodstock.


DNA techniques
Whole genomic DNA was extracted from alcohol-preserved fin tissue by means of a sodium chloride extraction of proteins followed by ispropylic alcohol precipitation of DNA (Miller et al. 1988). The polymerase chain reaction (PCR) was used to amplify a segment of the mitochondrial genome containing 188 base pairs (bp) of the O. mykiss control region and 5 bp of the adjacent phenylalanine tRNA gene using primers S-phe (5′-TAGTTAAGCTACG-3′) and P2 (5′-TGTTAAACCCCTAAACCAG-3′) (Nielsen et al. 1994).
Nomenclature for mtDNA control region haplotypes follow those given in Nielsen et al. (1997a, 1998). Amplifications were conducted in a total volume of 50 µL containing 1× retype (ST1, Nielsen et al. 1994; details in Results), suggesting either that they descended from a monomorphic population, that the population became fixed for haplotype ST1 during establishment and colonization, or that not all population haplotypes were represented in our sample. We thus developed an ad hoc model to evaluate the likelihood of ending with an all-ST1 sample given that the stock of origin was nonmonomorphic.
We consider three processes that, starting with a nonmonomorphic maternal stock, could lead to an all-ST1 sample: the sampling of females from the donor population that produced the eggs imported (founder effect), the mortality between eggs and reproductive fish contributing to establish the new stock (postfounding drift), and the chance of missing population haplotypes during our sampling process (sampling effect).
Each of these three processes can be viewed as sampling from a finite population, which is most properly modeled by a hypergeometric distribution. For the sample sizes and probabilities used in our analysis, the binomial distribution approximates the hypergeometric well. We therefore opted for computational simplicity and modeled the foundation of Santa Cruz populations as a chain of three binomial processes.
The number of donor females, different females that could have contributed to the Santa Cruz River stock, F, is calculated as 1    F=F/fec where E is the number of eggs imported and “fec” are putative values for average female fecundity. Assuming that the maternal females were randomly drawn from a particular population, we modeled the number of ST1 eggs effectively extracted from it and imported into Argentina, E ST1, as a binomial process:

2 E ST1 ≈ fec · Bin(F, φ) where φ is the frequency of the ST1 haplotype in the original population. The post-introduction mortality from eggs to founding fish, W, i.e. fish that effectively contributed to the Santa Cruz stock, is simply modeled as
3 W = surv ·E where “surv” are putative values of survival rate from eggs to founding fish. The number of ST1 fish in this founding stock is
4 where EST1/E is the proportion of ST1 eggs effectively imported as modeled in eq. 2. The number of ST1 fish in the sample taken from the present population (SST1) is
5 where n is the sample size and WST1/W is the proportion of ST1 individuals among the founding fish. It is assumed that the frequency of ST1 currently observed in the population is well represented by that of the founding fish. In other words, we assumed that there was a single, primeval bottleneck associated with initial establishment, after which the population expanded rapidly enough for the frequency of ST1 to remain reasonably unchanged. The probability of obtaining and all-ST1 sample from the present population is6 Finally, for given founding stock (φ is the frequency of ST1 in the maternal population), average fecundity (i.e., or number of donor females (eq. 1)), egg to founding fish survival (i.e., or number of founding wild fish (eq. 3), and sample size (n), the probability of obtaining an all-ST1 sample is given by integrating eq. 6 over all possible outcomes of eqs. 4 and 2:
7    The number of eggs imported, E, was set to 75 000. We used an array of values for “fec” between 500 (low fecundity) and 4500 (high fecundity), considering 2800 to be an average fecundity for typical Sacramento River rainbow trout stocks (Carlander 1969). These values correspond to a range of 17–150 donor females. We used values of φ consistent with the frequency of haplotype ST1 in different candidate donor populations of Santa Cruz River fish (Table 1).

We used values of “surv” between 0.00006 and 0.0029, corresponding to founding population sizes of 5 (very low survival) to 215 fish (high survival). Finally, we used a sample size n of 20, the number of wild fish sequenced in this study.
We did not consider in our model the chance of missing low-frequency population haplotypes during our sampling process. While this probability may not be unimportant for sample sizes of less than 10 individuals and frequencies of 0.85, it becomes low for sample sizes of 20 individuals. We therefore preferred to accept a small bias and avoid the need for the much more intensive calculations demanded by including three nested conditional probabilities in our model.

Stock Life history ecotype Halotype Halotype frequency (%)
MCCloud redband trout Resident (anadromous ancestry ST1 100
Northern California
Eel and Sacramento rivers Coastal Steelhead ST1 83
ST3 9
ST5 4
ST8 4
Central California coast
Russian River Steelhead ST3 41
ST1 40
ST5 15
ST8 4
Upper Sacramento River Steelhead ST3 62
ST1 21
ST2 14
ST5 3
Resident STH3 60

Note: Only those populations regarded as the putative parental sources for Santa Cruz River rainbow trout are included. Frequwncy data taken from Neilson (1996) and Neilson et al. (1994).
Results
Sequence data revealed that all Santa Cruz River fish, both anadromous and resident, had the ST1 haplotype described by Nielsen et al. (1994). Hatchery fish, on the other hand, were genetically different from wild fish. Only one of the five fish examined had haplotype ST1, while the remaining four fish had haplotypes ST3 and ST9 in similar proportions. Each of these haplotypes differed by only a single transitional base change from haplotype ST1 (G → A) at positions 1109 (ST3) and 1147 (ST9). All of these mtDNA haplotypes were previously reported by Nielsen et al. (1994, 1997b, 1998) and Bagley and Gall (1998) in rainbow trout populations from California and by McCusker et al. (2000) in populations from British Columbia.
Mitochondrial DNA haplotype ST1 is dominant in steel-head populations from the Sacramento and Eel rivers in northern California but among the putative parental stocks was found to be monomorphic only in the McCloud River rainbow trout (Table 1) and in the Río Santo Domingo rainbow trout populations from Baja California (Nielsen et al.1997b, 1998, 1999). We discard this last stock as a candidate source of Patagonian fish because Baja California trout did not contribute to fish culture at the time of the introductions.
Haplotype ST3 is rare in steelhead populations from northern California but is common in coastal populations from the San Francisco Bay area and dominant in resident populations from the upper Sacramento River and the Kern and Little Kern rivers (Nielsen et al. 1997b, 1998; Bagley and Gall 1998). Haplotypes ST1 and ST3 were found inequal frequencies in steelhead populations from central California (Table 1) (Nielsen 1996). Haplotype ST9 is rare (<2 percent) in coastal populations from California (J. Nielsen, US Geological Survey, Alaska Biological Science Center, 1011 East Tudor Road, Anchorage, AK 99503, USA, personal communication), but it is more common in inland steelhead populations from the Snake River in Idaho (Kucera andArmstrong 2001) and in inland populations from the upper Columbia River in Canada (McCusker et al. 2000).
To explore the hypothesis that a monomorphic sample of Santa Cruz wild fish could have originated through haplotype loss and sampling bias, as opposed to a truly monomorphic origin in the McCloud River, we applied our probabilistic model to two extreme alternative scenarios: a central California type parental stock, with a minimum 40 percent frequency of the ST1 haplotype, and a northern California type stock, with a maximum of 83 percent frequency of ST1 (Figs. 2a and 2b, respectively).
As expected from first principles of a binomial sampling process, the probability of haplotype fixation increases as the number of donor females and founding fish considered, indicating that it is highly unlikely that Santa Cruz River fish originated from such a stock (Fig. 2a).
When a northern California type parental stock is considered, results are less clearcut, with probabilities ranging between 3 percent and 45 percent depending on the values chosen for decreases (lower left in Figs. 2a and 2b).
When a central California type parental stock is considered, the probability of an all-ST1 sample remains very low (<1 percent) for practically all values of donor females and founding fish average fecundity and initial survival (Fig. 2b). This led us to scrutinize these parameters in more detail. Individual rainbow trout can have fecundities as low as 500 and as high as 13,000 eggs (Carlander 1969).
We used a range of 500–4500 to accommodate probable values for individual mothers of Santa Cruz river fish, but average fecundities reported for Californian wild populations are closer to the lower half of this range. For example, Hallock et al. (1961) reported a mean number of 2808 eggs for larger Sacramento River stripped females. Perhaps more relevant to our case, Wales (1939) reported that female rainbow trout trapped at Greens Creek, the trapping site of the first egg-taking station of rainbow trout on the McCloud River, weighed 2lb on average, with a mean fecundity between 1000 and 2000. Average fecundities lower than 2500 (at least 30 donor females) result in a probability of sampling only ST1 fish of less than 10 percent (Fig. 2), unless the survival from eggs to founding fish was very low (<0.00033 or <25 founding fish), in which case this probability becomes greater.
In summary, unless a small number of particularly large females (<17) had been used to produce the eggs shipped to the Santa Cruz River and (or) a very small proportion of the imported eggs survived to become founding fish (<25 individuals), the probability of obtaining an all-ST1 sample of 20 individuals from a northern California type parental stock is less than 10 percent (Fig. 2).
Discussion
Our analyses allowed us to establish the most likely origin of Santa Cruz River main-stem fish as well as to advance our general knowledge on the relative contribution of different parental stocks to rainbow trout in Patagonia. As previously suggested by microsatellite analyses (Pascual et al.2001), mtDNA data reinforce the idea that anadromous and nonanadromous Santa Cruz fish do not constitute independent lineages but have a common ancestry.
Wild fish are clearly differentiated from hatchery Danish stocks widely propagated in the region after 1950, providing strong evidence for an origin of Santa Cruz populations in Californian rainbow trout imported to Argentina during the first decade of the twentieth century. Additionally, these results indicate that the introgression from hatchery fish into the wild population has not been significant.
Although it has been widely accepted that early transplants of rainbow trout from the United States to locations around the world, including Argentina, came from the McCloud River (Scott et al. 1978; Busack and Gall 1980; Pascual et al. 2002a), historical records alone were insufficient to verify this, conferring some credence to the idea that other locations in northern California and southern Oregon could have potentially contributed fish to these early transplants (Behnke 2002). The fact that Santa Cruz River wild fish analyzed were monomorphic for haplotype ST1, together with the results from our probabilistic model, provides additional support to the view that the source of these populations was indeed the McCloud River.
It must be noted, however, that our approach does not consider some complex scenarios that could muddle the identification of parental stocks. First, Santa Cruz fish could have been derived from a mixture of fish from the McCloud and other northern California locations, leading to a larger probability of haplotype fixation than purported by our scenarios. Second, the founding population could have experienced multiple bottleneck events instead of the single event at the onset of the introduction that we modeled, increasing as well the probability of haplotype fixation. Since there are no conceivable bounds on the exercise of conjecturing combinations of parental stocks or bottleneck sequences, we did not attempt additional analyses, leaving it simply as a precautionary note.
The clear differences found between wild and hatchery Santa Cruz fish point at mtDNA analysis as a powerful tool to elucidate the ancestral genetic makeup of rainbow trout in Patagonia at a regional scale and to determine the relative contribution of stocks used before and after 1950. The occurrence of ST3 in hatchery stocks may suggest a Californian origin, most likely a genetic heritage derived from the Sacramento River rainbow trout, while haplotype ST9,which is rare in Californian wild stocks, hints at a complex ancestry of stocks imported from Europe, with probable contributions of non-Californian fish (e.g., British Columbia, where ST9 is more frequent). However, we analyzed only a handful of hatchery fish and larger samples from different hatchery stocks will be required to fully characterize the genetic makeup of this stock.
At present, these data make no suggestion as to what extent anadromy and residency in the Santa Cruz River are merely recreating the preexistent variation or have been modified in response to the specific selective pressures of the novel environment. Nevertheless, the identification of the genetic roots of Santa Cruz fish provides relevant background information to guide future research about the origin of life history variation in this river.
Regardless of the specific processes underlying life history variation, our results indicate that the Santa Cruz River may well constitute a unique, secluded reservoir of those ancestral McCloud fish widely distributed around the world during the late nineteenth and early twentieth century, which in their native range have been substantially affected by habitat modification and introgression from hatchery stocks (Busby et al. 1996; Nielsen et al. 1999; McEwan 2001).    ■

Oxygenation technology poised to transform aquaculture worldwide

by Stefan Dullstein, Head of Industrial Segment Aquaculture & Water Treatment, Linde Gases Division, Germany

Interest is burgeoning in a unique new low-pressure oxygenation system that is poised to transform the world of aquaculture.
The uniqueness of the technology is based on its ability to perform three critical functions in one system— dissolving oxygen in the water, producing the correct hydrodynamics and stripping out potentially harmful inert gases like nitrogen — via a very low energy requirement. Moreover, the system is easily installed, including as a retrofit to existing fish tanks, and is virtually maintenance-free.
SOLVOX® OxyStream, developed by Linde Gas and launched in August 2011, has been proven to improve the living conditions of fish inside the tank, allowing for a significant increase in fish production volume, optimise fish meat quality and considerably improve operations from an environmental standpoint.
Aquaculture, also known as aquafarming, is the discipline of commercially farming aquatic organisms such as fish, crustaceans, molluscs and aquatic plants.. Aquaculture involves cultivating freshwater and saltwater populations under controlled conditions – in contrast to commercial fishing, which is the harvesting of wild marine fish.
The Linde technology has been developed in response to a progressive trend that is seeing aquaculture production being transferred from sea cages to land-based sites for the full duration of a marine fish’s lifecycle. This change has confronted the industry with the challenge of oxygenating large fish tanks that can accommodate fish stock from infancy to maturity.
The ground-breaking patented SOLVOX® OxyStream is a combined oxygenation and flow system which not only dissolves the optimal amount of oxygen in the inlet water flow, but also distributes it evenly at an adjustable flow pattern through the tank, ensuring fish stock benefit from the physical exercise involved in swimming against the current. The flow regime can be fully tailored according to fish size, stock density and fish species, such as salmon or cod. The system comprises a standalone unit and is installed individually in each tank, so water flow and oxygen dosing can be individually controlled for each tank.
The micro-bubbles created by SOLVOX® OxyStream have the additional benefit of helping to reduce the concentration of dissolved inert gases such as nitrogen or argon. In particular, oversaturation of nitrogen, even in relatively small quantities, can endanger the wellbeing of fish stock, slowing growth and increasing the possibility of disease, and ultimately, even mortality. With the installation of SOLVOX® OxyStream, external degassing units to prevent inert gas build-up will, in many cases, become obsolete.
Depending on the application, pumping pressures as low as 0.05 to 0.2 bar are normally sufficient to oxygenate the incoming water, strip nitrogen and create optimal tank hydrodynamics. This low operating pressure makes the system very energy efficient. OxyStream also requires very low maintenance, because it is not associated with any ancillary equipment to manage water pressure.


Producing fish in captivity
Aquaculture using sea cages came into its own as recently as the 1980s, when the fish industry recognised it would be more cost effective to produce fish in captivity in the ocean rather than to trawl for wild fish. Today aquaculture is moving to on-land farming, with the most significant inroads being made in Norway, where there is a massive demand for salmon and cod. This places an enormous burden on farmers to produce fish more efficiently and cost effectively.
The limitation with a conventional on-land tank is the amount of oxygen available to the fish. Water can only provide a certain amount of oxygen, which is quickly respired, so there is a need to provide an additional source of oxygen.
SOLVOX® OxyStream is able to efficiently oxygenate sea water and can additionally be used during the fresh water phase of salmon. This allows fish farmers to operate a single oxygenation system in large tanks which can run on both fresh water and sea water. This significantly reduces costs compared with running separate saline and fresh water oxygenation systems.
This capability was recently demonstrated during trials conducted at Marine Harvest in Norway, the world’s largest salmon producer. Results showed that SOLVOX® OxyStream was the only oxygenation source suitable for rearing young salmon hatched in tanks running on fresh water, before gradually transitioning them to sea water. This creates an optimum environment in which to rear salmon, ensuring the correct oxygen levels throughout the entire production period and keeping fish stress levels to an absolute minimum.
By precisely predicting flow velocity SOLVOX® OxyStream is able to adjust this velocity in the circular on-land tanks. The fish, depending on their age and size, need a certain water velocity for optimum growth conditions, so  if the  velocity is incorrect, the fish won’t exercise, so it’s important to apply the correct water velocity in each application to keep them ‘working out’ against a robust current.


Parallel development
Typically, aquaculture farmers have grown salmon from eggs to about 100g in weight in small to mid-sized tanks. Once the fish are acclimatised to sea water conditions, they are usually transferred to sea cages.
This method has been constrained by some major challenges as there is always the potential for break-outs, allowing valuable stock to escape, while the high density of fish in this natural environment has the potential to foster diseases. So it makes sense to govern the entire lifecycle on land, where the health of the fish and the environment can be managed with more control.
Although the method was mooted as far back as ten years ago, at that time the costs were prohibitive owing to the high energy required to pump water through the dissolvers operated at higher pressure into land tanks and dissolve oxygen in the water. Today SOLVOX® OxyStream, with its unprecedented low energy demand, makes this possible.
Furthermore, new legislation imminent in Norway will allow farmers to develop fish to a size of 1000g on land, meaning a whole new market has opened for us. Research is already underway to investigate the on-land development of fish up to 4 to 5kg, harnessing our technology.
The largest tank equipped so far with SOLVOX® OxyStream is 15m in diameter, but aquaculture farmers are looking to increase the size of tanks – up to around 20 to 25m in diameter.
Immense interest
The industry has responded to the introduction of this system with immense interest and the Linde team currently has about 20 units piloting at customer sites, with many more proposals out there. It has fielded enquiries from North America, the UK, Norway, France and even from Saudi Arabia.
With interest levels so high, Linde has had to accelerate its activities to meet customer demand. As a customised solution, it is not a simple matter of ‘plug and play’. Each customer application needs to be approached from a unique perspective, needing professional, tailored input.
To showcase its technology in action, Linde is constructing a brand new test centre in Norway where customers will be able to see SOLVOX® OxyStream in action as it operates in windowed tanks. Training and equipment testing will also be carried out using the new centre as a base.
Environment
With the ocean’s reserves of fish steadily depleting and the demand for fish products on the increase, adding more sea cages along the world’s coastlines is not an adequate solution.
In this scenario not all the fish feed deployed to the sea cages is converted to fish meat — there is a lot of wastage. At the same time, a large amount of fish excrement released in a specific area tends to over-fertilise the ecosystem.
By comparison, the producing from infancy to maturity in on-land tanks has significant environmental benefits. The water flowing through fish tanks can be recycled to a high extent with excrement and feed residues filtered out effectively. As such, pollution to the marine environment and the possible spread of disease to wild fish is prevented.
Diseases can be properly handled in land based tanks. Fish farmers using sea cages may need to chemically treat the fish to get rid of sea-lice and this is prejudicial both to the environment and to the fish themselves. Sea-lice are not an issue in recirculation tanks, as these parasites cannot get into the system in the first place.
There are other advantages. Fish bred in aquaculture are, in many cases, genetically different to wild fish, so when there is a break-out and fish escape into the ocean, this could have an impact on the genetics of natural fish population.
A technology like SOLVOX® OxyStream was unquestionably needed to ensure the future success of the growing land-based aquaculture industry.  However, it’s not the end of the story. Although it is the most advanced equipment available to this industry at this time, Linde is convinced that it can improve the technology even further and research and development will be ongoing to maintain their position as a leader in the field.
The future
Envisaging a bright future for SOLVOX® OxyStream, Linde believes the vigorous interest being shown by aquaculture farmers is just the tip of the iceberg.
Norway’s fish farming industry, which is the most industrialised in the world, but by no means the biggest, produces about one million tons of salmon a year, but global tonnage – including all fish species, molluscs, croustades, and others, is as high as 50m tons per year. The largest aquaculture industry is in China, which produces about 70 percent of the world’s farmed fish.
However, the industry in China tends to comprise many small, family owned companies, using a low level of technology. Therefore, from a cost perspective alone, sooner or later China will begin to industrialise its aquaculture industry.    ■

Transfering Vitamin C from fish to embryos

Beneficial effects of ascorbic acid supplementation to broodstock of a select aquaculture species is well documented. At the present levels of feeding, dietary means of vitamin C does not meet the requirements for maturation, reproduction and needs of early life stages of larvae.
In addition, this nutrient is water soluble and readily gets accumulated by other organs before reaching the ovary.  For practical reasons, it is not possible to attain the desired level of a nutrient by conventional methods, hence innovative approaches are needed.   Mass transfer of nutrients via injection into broodstock is a novel method.
Two routes of maternal transfer of vitamin C in mature channel catfish (Ictalurus punctatus) prior to hormone-induced spawning were explored as a strategy to incorporate the vitamin and to determine its effect on reproduction and progeny performance.
The results of this study suggest injecting vitamin C prior to hormone-induce spawning, invokes transfer to eggs, improves reproductive performance, and may subsequently improve ontogeny performance.


However, the effect of vitamin C diminished with age and also in more natural conditions.  Our goal was to achieve predictable fish production of robust quality for healthy, efficient, higher surviving and able to adapt to common stressors and pathogens.
Improvements can be made in this area by new knowledge-based advances in nutrient delivery systems that may create large improvements in terms of production, feed conversion and survival in aquaculture production.

     
 Table 1.  Mass transfer of Vitamin C from broodfish to eggs/embryos
Species

Mass trasfer strategy

Dose

Reference

Rainbow trout

Oncorhynchus mykiss

Diet

1000 mg/Kg

Sandness et al. (1984)

Tilapia

Oreochromis mossambicus

Diet

1250 mg/Kg

Soliman et al. (1986)

Atlantic Cod

Gadus morhua

Diet

500 mg/Kg

Mangor-Jensen et al. (1994)

Rainbow trout

Oncorhynchus mykiss

Diet

500 mg/Kg

Blom and Dabrowski (1995)

Channel catfish

Ictalurus punctatus

Diet

500 mg/Kg

Zuberi et al. (2011)

Rainbow trout

Oncorhynchus mykiss

Immersion

1000 mg/L

Falhatkar et al. (2006)

Rainbow trout

Oncorhynchus mykiss

Immersion and Diet

1000 and 500 mg/Kg

Falhatkar el.al. (2011)

Japanese eel

Anguilla japonica

Injecting broodfish

50 mg/Kg

Yoshikawa, 1998

Japanese eel

Anguilla japonica

Injecting broodfish

1mL* /Kg

Furuita et al. 2009

Channel catfish

Ictalurus punctatus

Injecting broodfish

1 mL*/Kg

Chatakondi et al. 2010

                   *Vitamin emulsion was prepared by dissolving  Sodium Ascorbate in  0.9% NaCl Solution

Importance of vitamin C
Ascorbic acid is an essential micronutrient in the diet of teleost fish, which do not have gulonolactone oxidase activity. Vitamin C is needed for post-translatory hydroxylation of proline and lysine moieties in collagen, mineral metabolism to improve stress response and immunity, detoxification reactions, steroid synthesis and vitellogenesis.
Egg ascorbic acid deposition levels may easily be tailored by feeding broodfish with elevated levels of ascorbic acid before and after vitellogenesis. The accumulation of essential nutrients in eggs is dependent on the nutrient reserves in the female fish and therefore on the dietary intake of broodfish in the period preceeding and during gametogenesis. Hence, broodfish nutrition consisting of essential nutrients is important.
The earliest steps in embryonic development are dependent on and driven by maternal factors deposited in the oocyte during oogenesis.  Maternal factors are stored in the form of specific mRNAs, proteins, hormones and other biomolecules.  At egg activation and fertilization, these factors become available for embryogenesis, sometimes after a process of activation involving translation or protein modification.
It has been documented that vitamin C or ascorbic acid deficiency in larval fish has been associated with hyperplasia of collagen and cartilage, scoliosis, lordosis, internal hemorrhages, resorbed opercules and abnormal support cartilage in gills, spine and fins with deformities of the jaw and snout.


Based on recent research, vitamin C needs for reproduction and early life stages of fish are 10 times the recommended dose for raising young adult fish. These high levels cannot be met by dietary administration to broodfish because the nutrient is water soluble and readily absorbed / utilized by other organs during oocyte development.
It has been demonstrated in several species that nutrients in broodfish diet are transferred to oocytes through uptake of extra-ovarian substances from the maternal blood. Also, there was up to a 82.4 percent loss of ascorbic acid of the prepared commercial diet. A 3.8 – 8.3-fold increase of vitamin C in the diet generally results in 56 to 71.9 percent increase of total ascorbic acid in the eggs respectively. The fry produced from parents fed with elevated levels of vitamin C tend to have higher growth performance as compared with control groups.   Thus, there is a need in enhance ascorbic acid in the  broodfish. A diet with vitamin C content adequate for normal growth may not be sufficient for broodfish when the goal is to transfer ascorbic acids to embryos.
Reproduction and arval performance
Broodfish diet has a major influence on fecundity and egg quality. It has also been demonstrated that the nutritional status of broodfish can affect offspring quality. The accumulation of essential nutrients in eggs is dependent on 1) the nutrient reserves in the female fish and 2) the dietary intake preceding gonadogenesis. Vitamin C is needed for maturation, reproduction and larval metamorphosis. Beneficial effects include increased fertility, fecundity and egg quality.
Nutrients in broodfish diet are transferred to oocytes through uptake of extra-ovarian substances from the maternal blood. Immersion enrichment of eggs is another approach to introduce compounds and nutrients into eggs. Immersion enrichment followed by feeding fry with vitamin C enhanced feed was also found to be an effective method.
Injecting vitamin C in to broodfish during artificial- induced maturation improved reproduction and progeny performance. Efforts are underway to develop procedures to effectively and stably accumulate vitamin C in eggs by broodstock injections (Table 1).

Vitamin C needs of channel catfish 
Channel catfish is the single largest aquaculture fish species cultured in the United States.  Based on 2011 data, approximately 335 million pounds of catfish were processed, a reduction of over 50 percent compared to the best production in 2003 (2012 USDA NASS). The industry is currently struggling to keep pace with the increasing cost of feed, fuel, production inefficiencies, foreign imports and economy. Adopting hybrid catfish (channel catfish female x blue catfish, I. furcatus male), hybridisation can be used to improve productivity immediately by producing fish that exhibit hybrid vigor.
Based on numerous laboratory and field trials, hybrid catfish are superior in growth rate, feed conversion, survival, seinability and processing traits compared to commonly raised channel catfish.
A decade ago, producing commercial quantities of hybrid catfish was believed to be unattainable. Natural hybridisation is rare and artificial spawning of channel catfish has been historically low and with no effective ovulating agents available.
However, in the last 10 years, consistent and marked improvements were made in all the phases of artificial spawning and the hatchery production of hybrid catfish embryos. Improved production and consistent superior performance of hybrid catfish in commercial earthen production ponds has rejuvenated the industry with unprecedented optimism. Presently, a third of catfish farmers raise hybrid catfish in production ponds and hybrid catfish account for approximately 25 percent of all the total catfish processed in 2011.
Our goal is to achieve predictable fish production of robust quality for healthy, fast growing, survival and adapt to common stressors and pathogens and to varying environmental conditions. Improvements can be made in this area by new knowledge-based advances in live food production or nutrient delivery systems that may create large improvements in terms of production, survival and processing yield.


Preliminary findings
Broodstock preparation is the primary requisite for hormone-induced spawning of channel catfish in the production of channel x blue hybrid catfish. Hence, broodfish management techniques must be geared towards attaining maximum production of high-quality eggs and larvae because variable egg quality is one of the limiting factors in fish hatcheries.  Broodstock diet has been considered as one of the factors affecting fecundity, egg, and larval quality in fish. The accumulation of essential nutrients in eggs are dependent on the nutrient reserves in the female fish, and consequently on the dietary nutrient input of broodstock in the period preceding gonadogenesis. When eggs absorb water, it is possible to introduce compounds such as vitamins and minerals into the eggs with the water solution before water hardening.
It was hypothesised that injecting female broodfish prior to hormone-induced spawning would result in mass transfer of nutrients to improve maturation, ovulation, and subsequent progeny performance. Preliminary studies confirmed accumulation of vitamin C in ovarian tissue and invoked a positive response to ovulation, fecundity and egg quality. Mass transfer of vitamin C to the eggs improved growth and reduced  mortalities following Edwardseilla ictaluri disease challenge.
It appears that mass transfer of vitamin C to eggs is attained by injecting broodfish prior to hormone-induced spawning to improve progeny performance.     ■