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VOL 11 ISSUE 1 January 2019

A D VA N C E S I N P R O C E S S I N G & F O R M U L AT I O N An Aquafeed.com publication

Awesome algae An interview with Therese Log Bergjord, CEO of Skretting Group Microalgae: a new source of EPA and DHA

Revolutionizing tuna farming

INSIDE Tackling the challenges of phosphorus discharge


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AQUAFEED

3 Volume 11 Issue 1 2019

A D V A N C E S I N P R O C E S S I N G & F O R M U L AT I O N

Contents

REVOLUTIONIZING TUNA FARMING

* Cover story

The major challenge in successfully rearing tuna lies in the fish feed. Japanese company Nissui and BĂźhler have together developed a tuna feed that will enable sustainable, economic farming.

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INTERVIEW Therese Log Bergjord Therese Log Bergjord, CEO of Skretting Group and member of the Nutreco Executive Committee, talks about the company and the industry.

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AWESOME ALGAE

AQUACULTURE’S IMPACT

The many benefits of algal extracts to the production chain can provide a new revolution in aquaculture.

Idaho achieves success in mitigating the problem of phosphorus discharge from fish farming.

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ZEROING OUT THE ENVIRONMENTAL IMPACTS OF AQUAFEED: THE VERAMARIS SOLUTION

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With the help of natural marine microalgae, Veramaris bypasses the traditional aquaculture food chain with a new technology to provide a source of EPA and DHA that does not impact marine life and enables aquaculture to grow. FEED MANAGEMENT THE IMPORTANCE OF PRECISE BIOMASS ESTIMATION IN THE FEEDING OF FARMED SHRIMP

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While check trays and scales can help, many farmers are looking past traditional manual methods to keep their overhead down and optimize their profits. These farmers are embracing recent technological advances in gathering and analyzing data. COLUMNS GREG LUTZ—TRENDS & DEVELOPMENTS

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Changing your feed? You might want to change your fish also. ALBERT TACON— Interesting nutrition studies in the Chinese literature. NEWS REVIEW ▪ ▪ ▪ ▪

Can algae-based salmon feed reduce sea lice infestations? Aller Aqua partners with insect factory High levels of mycotoxins detected across the U.S. Converting wastes with fungi

NEW ON THE MARKET ▪ Organic metal amino acid complex to optimize trace mineral nutrition ▪ A bioactive feed supplement from sugarcane ▪ Crunching scientific data for marketing and communication

CALENDAR OF EVENTS

To read previous issues in digital format or to order print copies, visit: http://www.aquafeed.com/publications/aquafeed-magazine/

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From the publisher As we embark on our 21st year, we thought we should mark our ‘coming of age’ with a new look table of contents—and more significantly, new members to our team. I am pleased to welcome Lucía Barreiro as our Associate Editor and Dr. Albert Tacon as our Technical Editor. Lucía comes to Aquafeed.com with a strong background in aquaculture with a B.S. in Marine Biology, M.S. in Aquaculture and practical experience in aquaculture production, hatchery management, research and project management. In addition to working on the Aquafeed magazine, Lucía is also our new newsletter editor. Albert is well known to almost everyone in the industry from his years at FAO as an aquafeed and nutrition expert and as an independent consultant. He is also a board member of the World Aquaculture Society. On a personal note, we have worked together on numerous projects over two decades, and I am pleased to welcome him to a formal role on the magazine.

Suzi Dominy

Index to advertisers

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Andritz Aquafeed Horizons 2019

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Asian Pacific Aquaculture 19

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Biomin

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Extru-Tech Famsun

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GFFC: 6th Global Feed and Food Congress

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Hatcheryfeed Idah

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Mixed Cereal . Mixed Feed . Veterinary

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Prairie AquaTech

BC

Skretting

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Sparos

8

Victam International

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Wenger Manufacturing

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Publisher & Executive editor

SUBSCRIBE

CONTACT US

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Editorial: editor@aquafeed.com

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Advertising: sales@aquafeed.com

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Technical feed consulting: consulting@aquafeed.com

Hatcheryfeed magazine

General enquiries: info@aquafeed.com

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Therese Log Bergjord is CEO of Skretting Group and a member of the Nutreco Executive Committee. She joined Skretting from Compass Group, where she was heading up the Nordic business. Her background is in finance and business administration. She has considerable expertise in the aquaculture industry, having previously acted as Vice President Commercial for Pan Fish and Global Sales Director for Skretting. Previously, Therese spent 16 years in the oil and gas industry in various leading positions within ConocoPhillips. Therese Log Bergjord talks to Aquafeed.com’s publisher and executive editor, Suzi Dominy, on completion of her first year in office.

Interview AQUAFEED Perhaps you could tell us a little about your journey to becoming CEO of one of the world’s largest aquafeed companies; you worked at Skretting before – what brought you back? TLB Well, I originally studied Finance and Marketing at the University of Stavanger. After my studies, I spent a

with Therese Log Bergjord number of years in the oil and gas industry in various commercial positions. I joined the aquaculture industry in 2003, with Pan Fish, a global salmon farmer that later became Marine Harvest. And you are right, I spent two years in Skretting in 2007, as the Global and Norwegian Sales Director. I left the business for some years to become a Managing

Director, as I became the country and Nordic MD for Compass Group, a British food and support services company. I have been working in three very different industries and I have learned a lot. When the previous CEO of Nutreco called me to ask if I wanted to come back to Skretting, I knew immediately that it was a big yes.


8 I have been involved with food, as a business for a number of years. I am proud to be part of an important industry that I believe have a purpose – and Skretting can play a role in the development of the industry. And our mission – Feeding the Future…isn’t it great?

AQUAFEED There is no doubt that the aquafeed industry is facing the potential of a future ingredient shortfall. What steps is Skretting taking to address this issue? TLB If we are to increase food production sustainably, and ensure that the rising world population has access to essential food, it’s important that we continue to develop alternative sources of sustainable raw materials in our fish and shrimp feeds. It is Skretting’s ambition to work primarily with ingredients that do not compete with human consumption; novel ingredients such as protein from insects, microbes and even waste streams offer increasingly viable solutions. We see great value in working with other partners in this work and putting our money where our mouth is - investing in startups

and innovative technologies to ensure future supply for the entire industry.

needs of the different species, we have greatly increased our flexibility.

World-class R&D at our Skretting Aquaculture Research Centre (ARC) ensures that we are taking a leading position to overcome any impending shortfalls, and in fact we have already developed feeds for salmon including no fish meal or fish oil, with equal performance. Flexibility and independence is what we strive for when it comes to raw materials.

So while we are investigating all raw materials available to us and developing new sources, we will continue to use marine ingredients so long as they are viable from both a sustainability and commercial sense. We are involved in a number of fishery improvement projects, and greatly value being part of collaborative organizations such as SeaBOS, GSI and SSP to ensure we are working towards a more sustainable future, together.

AQUAFEED Skretting is now producing fish meal-free diets. Utilizing plant-based materials to feed fish is arguably unnatural: everything that comes from the ocean is naturally fed by the ocean. Is Skretting doing anything to develop ocean-based feed ingredients? TLB Well, I’ve been told by our R&D Director many times that in fact, raw materials are actually about nutrients, the different components within them, rather than the source itself. It is our detailed R&D and understanding that enables us to think this way. This means that for us at Skretting, a raw material is simply the way the nutrients are packaged, and when we break those apart into the components, and take into account the

AQUAFEED R&D is a strong focus for the company. What are your research priorities going forward? TLB One of our top priorities is to even further understand the raw materials we have today, and as I mentioned, also find new ingredients. It took us 30 years of research to become independent from fish meal, and several more to find a fish oil replacement. Skretting is in fact the only feed producer with the knowledge to make diets completely free of fish. We also see that there is a lot we can


9 do with the raw materials we have, so we will also continue to investigate modification of raw materials to improve their nutritional value. Hand in hand with this research is our work to understand animal physiology even better. Aquaculture is still quite a young industry, and there is a lot of opportunity to come. Feed is of course very important, but in the new markets, we also see that the way we feed and the interaction between the feed and the other elements is just as important. Working with digital solutions and modelling is also high on our development agenda.

AQUAFEED Would you tell me more about the investment in Israeli startup, ViAqua; will the technology afford Skretting a means to deliver therapeutics in-feed, or is this a farm management tool? TLB We’re really excited to see the first results from ViAqua, who are developing the first orallyadministered treatment for shrimp that improves resistance to viral diseases, including White Spot Syndrome Virus (WSSV), and prevents viral epidemics.

AQUAFEED With active participation in organizations such as The Global Salmon Initiative (GSI), The Sustainable Shrimp Partnership (SSP) and the Seafood Business for Ocean Stewardship (SeaBOS), you clearly see the need for Skretting to take a leadership role in promoting a sustainable and responsible seafood chain. What

can feed companies in general do to help achieve this goal? TLB We all have a role to play, no matter what size in the value chain. Feed is one of the most important factors in aquaculture when it comes to sustainability, and as one of the world’s largest suppliers, we feel an important responsibility in this discussion. We have a very clear mission of Feeding the Future throughout all of Nutreco. A collaborative approach is essential, we’re all in this together and no one player can make significant change alone. We see through pre-competitive initiatives we can all get involved and make aquaculture the most productive and sustainable industry it can be.

AQUAFEED In the three decades that I have been involved with the aquafeed sector, I have seen Skretting grow exponentially – both organically and through acquisition. What is the size and scope of the company now, and where is it heading? is there a specific goal in terms of production, geographic reach or product development? TLB Skretting made the decision several years ago to diversify outside of the traditional markets. This has turned out to be a very good strategy for us, and we are proud to say that we have built great competence in the shrimp and tilapia nutrition sector to add to our existing knowledge in salmonid and marine species. A key strategy point for us now is to grow in Asia in the coming years. We are confident that our global experience and competence can ensure healthy

growth in new markets. Expanding further into these areas also ties well with our mission.

A key strategy point for us now is to grow in Asia in the coming years. We are confident that our global experience and competence can ensure healthy growth in new markets. Expanding further into these areas also ties well with our mission.

AQUAFEED While the story has been mostly one of expansion, you recently announced the planned closure of your plants in the U.K., citing over-capacity in the market. Was this as a result of Marine Harvest building their own feedmills, or is the market shrinking? TLB Our decision to cease production in the UK is all about the market overcapacity, and of course any new factory will have an impact on the market. Within Skretting, we have great infrastructure throughout Europe to support our customers in the UK and intend to continue to do so.

AQUAFEED What do you see as the most pressing issues for the development of aquaculture?


10 TLB It’s quite a challenge to get good understanding that it costs money to ensure sustainability and safety of the food we eat, particularly seafood. We must continue to push for development in a positive way to ensure the future of the industry, and working in a collaborative and innovative way throughout the value chain will ensure that no challenge is too great to overcome.

AQUAFEED And finally, where would you like to see aquaculture in general – and aquafeed in particular – in the next 10 years? TLB In the big picture, we would like to see people eating more farmed seafood. It’s one of the healthiest sources of protein we can eat as humans, and we must ensure that its

value is understood so we can continue to feed the future. It is a fact that aquaculture is much more resource efficient than many protein sources – we need to make people understand what that means. For us at Skretting, this includes the implementation of novel raw materials in the feed that do not compete with those for human consumption. Ten years is actually a relatively short amount of time. We know that there’s so much R&D needed to ensure we are optimizing both the raw materials and animal physiology but we’re well on our way on this journey. Even in the emerging markets we are seeing high levels of technology with many new developments. We would like to see aquaculture creating job

opportunities, and supporting local community development. As I mentioned before our goal is to feed the future. We are committed to improving aquaculture to secure its position as the key protein producer of the future, wherever that may take place around the world. We plan to continue to innovate – and radically innovate – through our ongoing and ever expanding research activities at Skretting ARC, and utilizing all the tools at our disposal, such as those developed by the innovation arm of Nutreco. In this way, I truly believe we will fulfil our mission.

AFΩ


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NEWS REVIEW Highlights of recent news from Aquafeed.com Sign up at Aquafeed.com for our free weekly newsletter for up-to-the-minute industry news

Can algae-based salmon feed reduce sea lice infestations?

Photo: Helge Skodvin / Nofima

“When you cultivate microalgae so densely, you need a lot of CO2,” said professor Hans Christian Eilertsen of UiT. “The smeltery gives off a lot of CO2, NOx gases and residual heat, which we utilize to cultivate the microalgae.” Eilertsen said that the algae population remains completely uncontaminated despite gases being pumped directly from the smeltery and into the algae tank.

Researchers at Nofima and UiT, The Arctic University of Norway, have been tasked by the Norwegian Seafood Research Fund (FHF) to investigate if algae-based salmon feed can reduce sea lice infestations. “We are testing whether sea lice infestations can be reduced if the fish are fed using selected Arctic microalgae, which are produced using residual heat and exhaust gas from a smeltery,” explained Nofima researcher Sten Siikavuopio and research director Ragnhild Dragøy Whitaker. “Based on growth data and observations in the nutrition, it is

believed that oxylipins from omega-3 work as deterrents to sea lice and other closely related organisms. Algae are primary producers of omega-3.”

Through controlled processing of algae and analysis of fish that have eaten on one of three types of feed, researchers are able to control test the effects of the microalgae. The three feed types contain algae oil, fish oil and plant oil.

In order to carry out the project, substantial amounts of biomass must first be produced from microalgae. The production takes place at Finnfjord Smelteverk, located in the municipality of Lenvik in Troms County, Norway. Alongside regular production of metal for the steel industry, the pre-industrial scale production of microalgae takes place in a reactor that holds up to 300,000 liters.

“While algae and fish oils are rich in marine omega-3, the plant oil contains fatty acids other than marine oils,” said Birthe Vang, Nofima researcher. “Hence, the feed containing plant oil produces other oxylipins that we don’t believe will provide the same protective effect. The research can thus give us answers as to whether it is the marine omega3 in itself, or something special about the algae oil, that causes the fish to


12 receive substances that function as a sort of “shield” against the lice.” It is still too early in the project period to conclude whether one feed type or the other works best. However, Eilertsen says that the salmon eat the feed comprised of algae biomass.

“The salmon eat it, and it seems as though they like it very much,” he explained. The UiT professor sees significant industry potential in utilizing algae oil as fish feed, alternatively as an additional ingredient.

“Considering the high omega-3 content in the algae, and the areas of land that are lost due to the production of for example soy beans, the industry should, in the long term, consider replacing soy with algae.”

Aller Aqua partners with insect factory

Aller Aqua, DTU Aqua, The Danish Technological Institute, Hanneman Engineering and Champost are partnering with the company Enorm to establish Denmark’s first industrial insect production unit. The goal of the project is to produce 30 tons of inspects per day within four years from now. The project has been granted 15.9 million DKK from the Ministry of Environment and Food Denmark. One of the outputs from the project will be insect meal, and Aller Aqua’s role will be to help develop and test

products suitable for trout farming. Should this succeed, insect meal could also be tested as an ingredient for feed for salmon and tilapia.

before commercial application. Research and trials will be carried out at Aller Aqua Research in Büsum, Germany.”

“Insect meal has the potential to be a valuable raw material in fish feed not only due to its high protein content,” stated Dr. Hanno Slawski, Group Research & Development Director for Aller Aqua. “Several trials have been carried out with the inclusion of insect meal to replace or partially replace fish meal in fish feed with promising results, but further research is needed

Besides producing insect meal for inclusion in fish feed, Enorm Biofactory will also produce both insect meal and oil for use in pork and poultry production, and eventually human consumption. In addition, the project focuses on achieving no-wasteproduction with optimal usage of all nutrients and resources.


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High levels of mycotoxins detected across the U.S. Harvest samples from across the U.S. that have been submitted to the Alltech 37+® mycotoxin analytical services laboratory in 2018 show high levels of mycotoxins, particularly deoxynivalenol (DON), zearalenone, fusaric acid, fumonisin and HT-2. “Mycotoxins thrive in changeable conditions, with lack of rain, excessive rainfall or, sometimes, one after the other causing a perfect storm of contamination,” said Alexandra Weaver, global technical support with the Alltech Mycotoxin Management team. “The extreme weather events that we’ve seen this year around the world have led to increased occurrence of mycotoxins in many countries.” Samples collected across the U.S. include corn grain containing multiple mycotoxins, with an average of 7.0 mycotoxins per sample — more than 3.9 mycotoxins more, on average, than what was seen during the same period in 2017. Mycotoxins in U.S. corn silage samples are also showing

trations of zearalenone have been greater in 2018 corn silage. Type A trichothecenes (T-2/HT-2 toxins group) have also been detected at a greater occurrence in 2018 corn silage, at 43 percent, compared to 21 percent in 2017. Fumonisins also remain a frequent cocontaminant. an increase in occurrence this year, with an average of 6.8 mycotoxins per sample, compared to the 4.6 on average during the same time period last year.

Mycotoxins are seldom found in isolation, and when multiple mycotoxins are consumed, they may have additive, or even synergistic, interactions that increase the overall risk to the animal’s performance and health. As a result, the animal may have a stronger response than what would be expected if it were only experiencing a single mycotoxin challenge. While the type B trichothecenes (DON group) harvests are similar from 2017 to 2018, the occurrence and concen-

“These mycotoxins can affect animal performance and health, due to lower feed intake, gut health, reproduction and immune response,” said Dr. Max Hawkins, nutritionist with the Alltech® Mycotoxin Management program. Testing feedstuffs and finished feeds is important for livestock producers to understand these risks.” The annual Alltech 37+® mycotoxin harvest analysis test provides an assessment of contaminants in feed ingredients and potential risks to livestock. Between labs located in Lexington, Kentucky, and Dunboyne, Ireland, the Alltech mycotoxin management program has tested more than 26,000 samples of animal feed, each searching for over 50 mycotoxins.


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Converting wastes with fungi a growing amount of waste - using fungi that convert residuals and wastes from companies and households to food, feed and biofuels.

Researchers at the University of Borås are developing methods to grow fungi on materials that would otherwise have become waste. The goal of the project Ways2Taste is to produce climate-smart materials, including a new source of protein. The university is home to a unique research environment that addresses two of the world's major environmental challenges - a lack of resources and

"We usually say there is no waste— only resources. But our knowledge is not enough to utilize these resources. Ways2Taste is a new way of dealing with what would otherwise be waste," said Mohammad Taherzadeh, project leader and Professor in Bioprocess Technology at the University of Borås. Another participant in the project is the Food & Nutrition Science department at Chalmers University of Technology, along with 18 regional companies. The project is funded by the European Regional Development Fund. The waste products used in

Ways2Taste come from bakeries, ice cream factories, breweries, feed companies and food stores. They are divided into three categories: from relatively pure, such as day-old bread from bakeries, to really dirty, such as slaughterhouse by-products and fertilizers. The researchers will investigate how, via different processes, fungi can be grown on the waste to produce a fungal biomass. From this, several different end products can be created, including food. In the longer term, the research group wants to build a fungi center in Borås. "We want to be able to work nationally to develop useful fungi and help companies to use fungi in their production," said Mohammad Taherzadeh.


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PEOPLE IN THE NEWS Joel Newman to retire as AFIA's President and CEO

Joel G. Newman announced his retirement as President and CEO The American Feed Industry Association in December. Newman will continue to serve as president and CEO of AFIA over the several months and will assist with the transition through the end of 2019.

Joel has led AFIA for the past 15 years, during which he has ushered in a landmark era of change. Among his many achievements, Newman championed an expanded global focus on the development of international food safety standards and opening more international trade opportunities for the industry. He has represented the U.S. animal feed industry on international issues, including serving on the International Feed Industry Federation’s Board of Directors, and previously as IFIF’s chairman, and as a member of the U.S. delegation to the Codex Alimentarius Commission.

Andy Sharpe Andy Sharpe has been appointed President and CEO of Bühler North America. Andy has more than 20 years’ experience with Bühler, most recently as President and CEO of Bühler Aeroglide from April 2015 to the present. Prior to joining Bühler, Sharpe served as business manager for the snack and cereal division of APV Baker in Peterborough, UK, where he marketed complete food processing lines.

Changes at BioMar Niels Alsted

Patrick Campbell

Håvard Jørgensen

Niels Alsted, Executive VP of Business Relations in BioMar, has retired after 45 years. He has held various positions in BioMar and has been part of the executive management team in BioMar Group where he contributed to opening new markets, such as Chile and China. He recently chaired the Board of FEFAC and was a member of both the GSI feed task force and IFFO RS.

Patrick Campbell has become the VP of the Salmon Division. He was previously MD of the Scottish business unit and has headed the global R&D function within BioMar Group. He has also been one of the main drivers establishing a solid BioMar business footprint in Australia. He has been working for BioMar since 1998, where he started as product developer.

Håvard Jørgensen is the new MD in Norway, where he has been promoted from a position as Global R&D Director. He has been heading the global R&D organization since 2015 and has a very solid insight in to the Norwegian industry through his R&D work within Salmon as well as through his work heading the Norwegian industry organization “Sjømatbedriftene”.


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PRODUCTS & SERVICES

NEW ON THE MARKET Organic metal amino acid complex to optimize trace mineral nutrition Zinpro Corporation has launched ProPath®, its most versatile Performance Minerals® innovation for the U.S. and Mexico to optimize trace mineral nutrition for multiple species. The ProPath family of products delivers benefits to animals and customers that include a highly concentrated mineral formulation, exceptional uptake and absorption, biological efficacy, diet flexibility and consistency, product stability, solubility and sustainability. ProPath is classified as a metal amino acid complex. It stands alone as the only organic performance trace mineral product that combines two specific amino acids, each bound (or complexed) to a mineral, such as zinc, copper, iron or manganese, in a

stable, 1:1 ratio. ProPath’s unique molecular structure results in a more sustainable choice to improve animal wellness and performance. “Our ProPath product line relies on two synergistic, amino-acid absorption pathways (cationic and anionic) to elevate trace mineral nutrition accessibility in the animal,” said Bill Scrimgeour, CEO, Zinpro Corporation. “By optimizing these pathways, our ProPath innovation advances the science of animal nutrition, allowing minerals to be more effectively absorbed and metabolically available in the animal’s organs, tissues and enzyme systems. With greater mineral optimization, ProPath provides multiple benefits – for the animal, the producer and the environment.”

ProPath is flexible for use in a wide variety of diets, including conventional or specialty, like non-GMO), wet or dry. https://www.zinpro.com

A bioactive feed supplement from sugarcane The threat of antimicrobial resistance in aquaculture is an ongoing challenge to the industry and general public health around the world. It has been estimated that more than 90% of bacteria originating in seawater are resistant to one or more commercial antibiotics. The greater concern is that of the 51 commonly used antibiotics in agriculture and aquaculture, 39 are also extremely important to human medicine. If resistance to

these antibiotics continues to increase in aquaculture, there will be devastating flow on effects to disease rates and production outcomes. For this reason organizations such as the World Health Organisation (WHO) are promoting the “One Health” approach. This recognizes that to address health in an industry like aquaculture, the implications to human health and the environment must be addressed at the same time. This has

lead to the rise in popularity of phytogenic feed additives as an alternative to antibiotics. Phytogenic feed additives are plant based feed additives that are designed to give a functional benefit to aquaculture species. They are often from underutilized agricultural streams and therefore, are a useful option in increasing the sustainability of the industry. Sugarcane


17 (Saccharum officinarum) is a source of these phytogenic feed additives and is well known to have antimicrobial properties, whilst also having some of the growth promoting effects of antibiotics in production animal systems. Extracts from sugarcane are a natural phytogenic feed ingredient not used in human medicine as an antibiotic, are an ideal candidate to replace antibiotic feeding in aquaculture. For this reason some commercially viable sugarcane extracts are now available on the market.

T0 Control

Polygain™ is a natural plant extract from Saccharum officinarum; it is polyphenol rich, high in antioxidants, rich in minerals, nutrients and essential amino acids. Polygain ™ also has antiinflammatory and antibacterial properties. Polygain™ is a registered trademark of The Product Makers (TPM) which holds numerous global patents on the production and properties of Polygain™. Aquaculture trials in Bangladesh (Chittagong Veterinary and Animal Sciences University, Department of Fisheries Resource Management) and in Australia (James Cook University Centre for Sustainable Tropical Fisheries and Aquaculture) using Polygain ™ as a feed supplement in doses from 0.2% to 0.6% (w/w) delivers beneficial results to the intensive farming of a wide range of aquatic species. These studies cover Tilapia (Oreochromis niloticus), Pangus (Pangasius hypophthalmus), River Prawns (Macrobrachium rosenbergii), and Black Tiger prawns (Penaeus monodon). Polygain™ tilapia growth trial

T1 0.2% w/w Polygain™

T2 0.4% w/w Polygain™

T3 0.6% w/w Polygain™


18 Overall, feeds containing Polygain™ as a supplement has been found to deliver the following benefits in a dose dependent manner: - Improves overall growth performance - Attracts fish to the feed

- Maintains water quality & environment - Improves feed conversion ratio - Antioxidant effect improves fish meat odor and texture - Up-regulates gut microbiome (balance of healthy gut bacteria)

Mechanism of Action Extensive pre-clinical trials on the bio-

active compounds of polyphenols and flavonoids from Polygain™ has resulted in the following postulated mechanism of action.

Shane Mitchell, CEO The Product Makers Australia Bioactive Division E: SMitchell@tpm.com.au


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Crunching scientific data for marketing and communication Research-based companies are generating data at an unprecedented rate. This has been particularly observed in the feed sector, where suppliers are developing technologies that improve culture conditions and productivity to maximize return on investment, and are creating innovative formulations and additives that address the challenges producers are facing. Common examples include research efforts to find alternative protein sources, feed additives to replace antibiotics, ingredients that contribute to decreasing the fishmeal level in aquafeeds, and natural additives that enhance feed intake, digestibility and growth performance. Overall, a multitude of alternative ingredients and additives are continuously entering the market. However, these products commonly require scientific evidence before being readily accepted by the market.

Too much data, too little time R&D departments are often focused on generating scientific data and writing reports, whereas Marketing and Commercial departments have the challenge of communicating scientific information to clients. Yet, communicating scientific data efficiently is time-consuming, and time is increasingly valuable and limited. The focus of scientists is to generate data that support innovations and product applications, while marketing and com-

the goal is to facilitate the communication between R&D, Commercial, and Marketing & Communication departments of research-based companies and catalyze the process of communicating scientific data to any audience.

mercial managers are focused on developing and applying tools to meet sales goals. As a consequence, communication materials are often outsourced to marketing agencies, a practice that often creates communication problems due to the lack of specialized scientific knowledge. Additionally, scientists and communication designers speak different technical languages that are often difficult to combine. Ultimately, this results in multiple back and forth interactions between marketeers, designers and scientists that are argely inefficient and time consuming.

Changing the game A new alternative to traditional marketing agencies is now available. Scite is a new service provider that crunches scientific data and breaks down complex concepts and results into simple and easy-to-use contents that can be rapidly assimilated. Ultimately,

Scite has a multi-disciplinary team of scientists, communication designers, and illustrators, who get the science right and have the skills to transmit a technically accurate message in simple terms. The starting point for each assignment can be a scientific report, a dataset, or even an idea. The final output is a tailor-made content in the format that best suits the clients’ needs, such as infographics, marketing materials, technical and scientific articles, among others. Scite’s goal is to fill the gap between the generation of scientific data and the delivery of communication outputs, thereby reducing the time companies need to crunch science into marketing contents. Ultimately, Scite safeguards that the capital-intensive generation of scientific data is fully exploited by its clients in the form of high impact outputs.

Miguel Costa Leal, Ph.D. General Manager Scite—Science Communication Lda E: miguel.leal@scite.pt www.scite.pt


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Aquaculture’s Impact Obstacle: Phosphorus discharge from aquaculture The race to put fish on the table is in lockstep with the world’s rising food demand

But fish farms have been criticized for producing concentrated levels of nitrogen and phosphorus, creating a growing source of nutrient pollution in waterways and oceans. Critics claim that rising levels of phosphorus and other pollutants from excess excrement, uneaten food, and other organic waste could plague the world’s waterways. Phosphorus is necessary for a number of metabolic functions in fish, such as bone development and scales, said Gary Fornshell, an aquaculture extension educator with the University of Idaho, but too much phosphorus can clog waterways with algae and excessive plant growth. Such eutrophication uses up oxygen in the water, suffocating aquatic life and creating impaired waterways. Fornshell uses Idaho’s Snake River Valley to explain how producers are meeting phosphorus discharge guidelines and maintaining the state’s healthy waterways. Idaho accounts

Photos: @John Mollison

The Food and Agriculture Organisation of the United Nations (FAO) reports that aquaculture grew about 4.5 percent in 2017, with a total harvest of 83.6 million metric tons.

for about 68 percent of all trout production in the United States, making it the nation’s largest commercial producer of rainbow trout. In the early 1990s, Idaho’s Department of Environmental Quality (DEQ) and federal Environmental Protection Agency (EPA) cited issues with excessive plant growth, finding the mid-Snake River water-quality limited. Fornshell explains that background levels of phosphorus already exist in the area, as well as nonpoint runoff from surrounding ag land and cities. At the time, “the DEQ stated that

point sources (such as aquaculture) were contributing 15 percent of the phosphorus waste,” he said, adding that point sources often are the target of emission guidelines because they offer identifiable levels of target pollutants and are regulated under the Clean Water Act. The “water-quality limited” designation triggered requirements under the Clean Water Act and a Total Maximum Daily Load (TMDL) was calculated, showing allowable levels of phosphorus in water being cycled through from flow-through aquaculture systems (raceways) to the Snake River or its tributaries.


22 “For the industry as a whole, the waste-load allocation mandated a 40 percent reduction of total phosphorus discharged from the baseline in the early ’90s,” Fornshell said, “so that waste-load allocation was put into producers’ permits.”

A Profitable Partnership Using feed formulations and new ingredients to improve standards

Solution: Producers come into compliance Years later, data shows that aquaculture producers have met that guideline and gone beyond it, Fornshell said, and those efforts to meet the TMDL came on several fronts.

First is oversight. Several agencies regulate the aquaculture industry in Idaho, according to the state DEQ website. In addition to the EPA and DEQ, commercial aquaculture facilities must be licensed by the Idaho State Department of Agriculture (ISDA) as well as have their effluent control facilities approved by DEQ. The Idaho Department of Water Resources (IDWR) oversees water rights for facilities, and the Department of Game and Fish and other agencies also play a role. “In terms of trout farming and flowthrough systems, Idaho has been at the forefront of oversight for responsible aquaculture,” Fornshell said.

Second is construction of aquaculture facilities. Idaho has cold-water operations for trout steelhead, salmon, and sturgeon, and warmwater facilities that raise catfish and tilapia. Whether operators use a set of ponds or earthen or concrete

Peter Fritsch, president of Rushing Waters Fisheries

Peter Fritsch, president of Rushing Waters Fisheries near Palmyra, Wis., has been seeing phosphorus discharge numbers go down and production numbers go up for most of his 21 years in aquaculture. “Feed producers have been working on better formulations for years,” Fritsch said. “Feed conversion rates have gotten better and better.” The Rushing Waters trout farm sits on 80 acres and has 56 flow-through ponds or earthen raceways and nine artesian wells. Water is discharged back to the natural waters of Wisconsin. “This business is about the slow process of raising good fish, but it’s fastpaced too because every day there are challenges,” Fritsch said. In 2015, the challenge seemed almost insurmountable. The state’s

raceways, situated in series or in parallel, all operations must be built to minimize the release of nutrients and solids to surface or groundwater and comply with permit requirements.

The third component addressed was diet. As Fornshell shared, phosphorus is an essential mineral for fish growth. But research has been ongoing to determine dietary requirements for fish farming.


23 Department of Natural Resources called for a point-source reduction in phosphorus discharge going from 1.0 part per million to 0.075. “That was a drastic reduction,” Fritsch said. “Now we didn’t have to hit it on Day 1, but we had to make strides quickly.” He said early meetings within the company led to a few options: reducing production levels; installing a waste-water treatment plant; buying phosphorus credits; and diet adjustments.

The first three would force Rushing Waters to take a big hit financially. “We dove into feed,” he said. “But I did not think that feed alone would get us there.” The company began testing a product from Prairie AquaTech in South Dakota, using its ME-PRO® plant-based protein ingredient with impressive results.

The final area addressed was wastetreatment systems. Fornshell said commercial operators have explored design options for settling basins to capture and remove solids before they enter receiving waters.

Results: Good numbers, but where to go from here?

In over two years, and increasing inclusion levels of ME-PRO, Rushing Waters lowered its phosphorus discharge 69 percent with indicators that the phosphorus levels will decline further. The fishery has always operated using best-management plan practices, Fritsch said, monitoring when to feed, how much to feed and buying highquality feed. But he says that the results from Prairie AquaTech’s product were more than he could have ever hoped for. “Without ME-PRO® there is no way we could have gotten to this level just on feed. We view it as a long-term partnership.” “First, researchers found that from a practical standpoint, at least 90 percent of all feed goes into production trout rather than fry and fingerlings, and their dietary needs for phosphorus are lower because they’re not as actively growing,” Fornshell said. Up until then, feeds were

As a result, aquaculture producers began using feeds that more closely matched the fish’s dietary requirement for phosphorus and the forms of phosphorus in the feeds are more digestible so that less phosphorus enters water systems.

formulated based on the dietary phosphorus requirements of fry and fingerlings. “And second, feed producers began looking at what forms of phosphorus are better utilized, so they began using forms that were much easier for fish to digest.”

Idaho’s success in mitigating phosphorus discharge is a product of coming at a problem from all angles. Going forward, however, Fornshell said, “In terms of what is practical and affordable, producers are close to being at the limit of what they can do. We really have to keep working to develop feed formulations where greater amounts of phosphorus are digested by fish to minimize what is excreted in the feces and urine.” He adds that the state currently is undergoing another TMDL revision. “There’s a concern that at the end of the day, they are going to ratchet down waste-load allocation even more.”

AFΩ More information John Mollison South Dakota Innovation Partners E: john@southdakotainnovation.com


The leading international technical conference series for aquafeed professionals Brought to you by Aquafeed.com

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12 AQUAFEED HORIZONS JUNE 12, 2019 l

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Charles Engrem Key leading indicator best practices in the extrusion/drying process

Tania De Wolf Early life nutrition – producing quality fry

Paul D. McKeithan IoP: Leveraging people, process, and platforms to maximize aquafeed production

Thomas Ellegard Mohr Extruded aquafeed quality management; relationship between technology and extruded aquafeed quality

Luís Conceição Modelling tools to evaluate aquafeeds

John Mollison Improving water quality through feed

Robert Strathman Optimal design and processing of aquafeed

Julian M. Foerster Insects as a substitute for fishmeal: Influence on the extrusion process and the product properties of carp feed

Daniel Stoffner Process optimization in aquafeed, a challange with fluctuation in raw material qualities and formulations

Dana Nelson Practical realities associated with micro pellet production

Nicola Tallarico Optimization of the use of lipids in

Ester Santigosa A microalgal oil containing EPA+DHA as a source of omega -3 in aquaculture species

aquaculture diets


25

Revolutionizing tuna farming

To meet global demand for bluefin tuna while protecting the species, many seafood processors rely on fish farms. The major challenge in successfully rearing tuna lies in the fish feed. Together with the Japanese company Nissui, Bühler has developed a tuna feed that will enable sustainable, economic farming.

Tsuyoshi Goto is on board a fishing boat owned by Kaneko Sangyo, a group company of Nissui Cooperation, one of the largest Japanese seafood processors. It’s 9 a. m., and the ship is near Kabashima Island in the Sea of Japan, about 90 kilometers from the mainland. It’s feeding time. The bluefin tuna are excited, swimming circles around the aquafarm. They know what is about to happen. They are fed three times a week in the spring. Goto-san bends down, reaches into the sack behind him, and takes out a handful of oblong pellets. They look like a mix of chocolate and energy bar, but they definitely smell fishy. “This is the future of tuna farming,” he says, tossing the pellets into the 40 -by-40-meter floating cage. This is the signal that feeding time is about to start. The handful of pellets is followed by a canon shot that blows one ton of feed into the cage. The tuna comes. Nearly 1.2 meters long, the creatures stir the water, racing for the energy bars. Tsuyoshi Goto is Plant and Quality Manager at Farm Choice, a subsidiary of Nissui. Farm Choice produces feed for fish farming. Together with Bühler, his team developed these extruded pellets, the key to sustainable, economical tuna farming.

Story and photos by Martin Hoffmann.


26 Bluefin tuna are endangered . Since the end of the 1990s, the population of blue-fin tuna maturing in the wild has decreased by 80 percent – an alarming trend. Yet, fish consumption has continued to grow for years. In 2016, 151 million tons of fish were consumed worldwide, corresponding

to over 20 kilograms per capita. Among the Japanese, fish takes the number one spot when it comes to most popular foods. On average, each Japanese citizen eats 66 kilograms a year, with this tendency growing. At the top of the menu – tuna. Of course,

this is also related to the increasing popularity of sushi across the globe. Demand for high-quality tuna has skyrocketed in recent years. The catch quotas set by the government are nowhere near enough to satisfy the hunger for sushi, resulting in illegal fishing. Many are unable to resist the appeal of earning a quick buck. At Japanese fish markets, a fully grown 200-kilogram tuna can easily bring in tens of thousands of dollars, with the record price for a single fish coming in at 1.3 million dollars. In 2013, a sushi restaurant purchased this high-cost bluefin tuna at the prestigious New Year auction at the Tsukiji fish market in Tokyo. Naturally, this exorbitant price gained the media’s attention. One thing this development makes clear is that tuna has become a multi-billion dollar business. But the consequences are as obvious as they are alarming. Bluefin tuna has become an endangered species. The huge sack filled with compound feed has done its job here and the crew make their way to the next cage. In total, there are around 1,600 of them spread around the Japanese islands.

Assessing quality at the fish auction

Kaneko, another subsidiary of the Nissui Cooperation and the link between Farm Choice as the feed producer and the Nissui brand, operates several farms in the Goto Island archipelago, no more than 20 kilometers from the harbor. In the morning, Okuura is always the first stop – it’s where the large tunas can be found. At four-years-old and weighing in at 80 to 100 kilograms,


27

Some 1,600 tuna cages dot the waters around the islands of Japan.

they are ready for harvesting. Kaneko manages the farms, is responsible for cutting the fish, and processes the tuna in several plants. The company sells cut-to-size pieces to sushi restaurants as well as end consumers around the world.

the trend in the ‘90s, testing various methods and resulting in the establishment of Kaneko and Farm Choice.

The concept of tuna farming is nothing new. Research in the area began as early as the 1970s, when it was first discovered that the population of bluefin tuna was declining while demand was growing. The research is now led by Japanese Kinki University. But Nissui also identified

The main problem in tuna farming is the survival rate. Less than 1 percent of fish eggs hatch. By comparison, the survival rate of other farmed fish, for example salmon, is between 20 and 40 percent. And more importantly of course, is that the few hatchlings quickly grow in terms of size and

The right feed

weight without major losses. The issue: tuna are predators. If they aren’t fed enough or at all, or are given the wrong feed, they are no stranger to cannibalism and end up depleting their own kind. On top of that, the bluefin tunas are very picky eaters and prefer to prey on herring and mackerels. Past experiments with artificial feed failed miserably. Consistency, the mixing ratio, and shape were all too complex to produce and the tuna sim-ply did not accept them. The only remaining option was to eat fresh fish.


28 Consider this example: around 2,000 tuna are bred in a single cage. Each week, they need 40 tons of mackerel and herring. In total, the fish spend four years in a floating cage. Over time, this equates to 8,320 tons of fresh fish that are fed per unit and cycle. And there are more than 1,600 cages like this in the Sea of Japan alone. This is not very efficient. However, it shows that the key issue lies in their feed. Tuna farming may earn money despite the high costs associated with purchasing feed, but this is mainly due to the extremely high retail price of the fish. This approach to production is far from sustainable. In parts of Japanese waters, even mackerel and herring are considered endangered species. There has to be another solution.

Collaboration with Bühler “In 2010, we decided to collaborate with Bühler. Bühler is the expert technology provider when it comes to fish feed production. For us, it was clear that we could come a long way with their expertise in extrusion,” says Goto-san. It is now evening. Goto-san has joined his colleagues for dinner, including Urs Wüst, Key Account Manager and fishfeed expert at Bühler. Having lived in Japan for 11 years, he is familiar with the culture and customs, and now provides Nissui with support on behalf of Bühler. What else is on the table other than sushi – a fitting meal after a day at sea. Of course, bluefin tuna is also a

Specialized tuna feed in production

must, sliced as sashimi or formed into a nigiri with wasabi and rice. What’s special about all this? The tuna served comes from one of the Kaneko fish farms. Fed exclusively with compound feed, produced on Bühler extruders.

Shortly after establishing contact in 2010, Goto-san and his team traveled to Switzerland to work with Bühler on the tuna feed production process. The main difficulties lie in the shape, composition, and different consisten-


29 cies required for the feed. The shell of the pellets has to be both solid and soft, yet pliable, while the inside has to be soft, but not liquid. This recipe calls for the Swiss experts. The lower the portion of fishmeal, the more sustainable and cost-effective the feed. “It contains all the ingredients that tuna need to grow: fishmeal, fish oil, vitamins, and minerals. Our feed enables healthy farming of tuna, without using antibiotics,” says Gotosan. After two years of intense collaboration, the pilot phase can begin. In 2012, Bühler installed three extruder lines in the Farm Choice production hall farm in Karatsu.

Developing the feed In addition to the standard feed with mackerel and herring, several units are also fed with the extruded compound feed. Thanks to the higher nutritional value compared to fresh fish, a much lower amount is required. Instead of the standard 40 tons per week across five feeding days in spring, now only three feedings of 4.5 tons are necessary a week, a total of 13 tons. This saves time, personnel, and above all, costs for boats and fuel. And the pellets are also easier to store. Unused feed can be put back and used at a later point in time. Yet another advantage over fresh fish, which has to be caught, portioned, frozen, transported, and finally thawed before the tuna can chow down. During the test phase, the feed is optimized in close collaboration with Bühler.

The three extruder lines in Karatsu have been working since 2012.

With underwater cameras in aquafarms, Goto-san studies the behavior of the fish during feeding. His team observes that the tuna hesitate before eating the pellets. Compared with real fish, the feed doesn’t have a “front.” Predators always eat their prey head first, as this makes them easier to swallow due to the direction of their scales and fins. As for the pellet, the tuna is unable to detect a head, which is cause for confusion. “So we changed the shape of the feed yet again. It was important to develop a kind of a point, sort of like a nose, to signalize to the tuna that this is the front,” explains Gotosan. Minor adjustment, major success. The fish accept the feed. This recipe forms the basis for further experiments, with the objective of continuing to reduce the fishmeal content by 50 percent. The lower this portion, the cheaper and above all more sustainable production becomes.

A toast to success Tsuyoshi Goto, his team, and Urs Wüst have just reached the sake. This rice wine goes perfectly with sushi. “I am confident that we will meet our goal. Tuna farming will turn into a huge business. Of course, it’s a risk, but I am certain that we will continue on our course for success,” says Wüst. But first, another toast to their success.

AFΩ

More information Urs Wuest Bühler Key Account Manager E: extrusion@buhlergroup.com


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INGREDIENT FOCUS

Awesome algae The Blue Economy is an important part of achieving zero hunger and the many benefits of algal extracts to the aquaculture production chain can provide a revolution in aquaculture.

Photo: Mark Yokoyama

By Gianluigi Negroni

Traditional disease control methods include the use of vaccines and antibiotics to prevent and control diseases. In recent years, public pressure against the use of antibiotics in farm animals shifted the focus from ‘killing pathogens’ to ‘promoting beneficial microbes’ or ‘bio-friendly agents’. This is a novel approach to improving gut health, growth performances and wellbeing of farmed aquatic animals. Several terrestrial animal probiotics are now used in aquaculture species.

But a good probiotic for aquaculture should be able to colonize, establish and multiply in the host gut - and not all commercially available products can survive and/or remain viable at optimum concentration in fish guts, due to their non-fish/marine origin.

Seaweed for animal immunity The effect of algal extract properties on the immune mechanism of aquatic

animals has been well studied in recent times, because of their capacity of linkage due to their complex branching structures in three dimensions. These properties reside in the branched heteropolysaccharides. They are not present in terrestrial plants. They are complex carbo-hydrates that influence the immune system via different pathways, some of them not yet well understood. Algal polysaccharides are complex forms that contain repetitive structural features that are chains of


32 mono-saccharide residues joined together by glycosidic bonds; thus, they form polymer (-type) structures represented in the form of chains (Olmix 2018, Aquaponics seminar). In higher organisms, including aquatic ones, the huge variability of polysaccharide structures has the potential to regulate mechanisms in different cell interactions. In particular, sulfate is very active in various biological processes, noted in polysaccharides extracted from marine macro-algae. Research by Olmix and INRA shows how green seaweed could be the future for aquaculture farming. In vitro research gave very interesting results with Ulva armoricana extract Marine Sulfate Polysaccharides (MPS) in Bretagne. The test was done with cellules from pig intestinal epithelium to see the effect of the MPS on different strains of bacteria. The results were very good with strong biocide activities on the main bacteria Gram negative and positive (E. Coli, Listeria monocitogens, S. aureus, Streptococcus sp, ‌ 42 strains) that negatively influence animal farming. The gram-positive bacteria were the most sensitive to the seaweed extract treatment. The exact antimicrobial mechanism is not yet understood and more studies are needed. Meanwhile it is assumed the MPS active in the intestinal epithelial cell that produce cytokines are the immunity answer against the bacterial pathogens. Moreover, the mucosal immunity is also increased, giving additional protection. Finally, the study confirms the other precedent studies where the MPS have immune system

modulating and anti-inflammatory properties in animal farming. Now in vivo tests trials are ongoing on in different aquaculture farmed animals to confirm the above.

Gut health another beneficial use of seaweed extract It is well known that aquatic animals are in contact with several pathogens in the water column. This is particularly accentuated in marine water where the aquatic organism needs to drink a large amount of water to balance its osmotic body regulation with the surrounding environment. The microbiome in fish gut microbiota contributes to digestion and can affect nutrition, growth, reproduction, overall population dynamics and vulnerability of the host fish to disease; therefore, this microbial community is highly relevant for aquaculture practice.

Moreover, many biosynthesis and metabolism pathways of carbohydrates, amino acids and lipids are significantly enhanced by the gut microbiome in some fish. These findings suggest that the metabolic role played by the gut microbiome in fish can be affected by feeding. At birth, the epithelial surfaces of fish are colonized by large numbers of microorganisms (microbiota) that form commensal or mutual relationships with their hosts. The digestive tract houses the majority of these microbes, where they influence a broad range of host biological

processes. All the above studies arrive at the conclusion that negative interactions of feeding can potentially greatly altered the gut ecosystem provoking an adverse effect on the digestive physiology with important loss in aquaculture animal performances. Seaweed extracts can beneficially interact with the gut microbiome resident commensal communities; this can provide positive performance, increase the feed conversion ratio, beneficially influence fish gut activity, increase growth performance, better develop the innate immunity, and optimize the functioning of the gastrointestinal system. The use of some seaweed extracts can enhance the length and strength of the intestinal villi and microvilli and enhance, maintain and increase the mucus protective cover. This can greatly increase the absorption and uptake of nutrients and energy of the feed. The good health status of mucus prevents opportunistic bacteria from attacking the gut and epithelial cell on the gut villi; they are rapidly removed from the gut areas, permitting the appropriate development of the commensal microbiome. Also the villi will be longer and together with the mucus layer will benefit the microbiome. All this provides healthy gut conditions for the modern aquaculture farm: feed cost reduction, improved animal health with less veterinary costs, shortened production cycle, better genetic animal expression, and good growth performances rates.


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ALGAE BROWN

RED

GREEN

Alginate, Fucoidan

Galactan, Carrageenan

Ulvan

Fucose, Xylose, Uronic acid, Galactose, Sulfate

Agalactose sulfate, 3,6-Anhydrogalactose

Rhamnose sulfate, Aldobiuronic acid sulfate

Kappa, Iota, Lambda types

Fig. 1. Main group of seaweeds and their extraction components (Olmix 2018).

Use of seaweed extracted Marine Sulfate Polysaccharides (MPS) on immunity Seaweed extracts are considered the future in several aquaculture areas. They are produced by mechanical separation or hydrolysis with consequent fractionation by ultrafiltration. The length of the molecules provides different biological characteristics of the algal product. The Brown seaweeds produce fucanes, alginate and uronic acid; the red seaweed galactan and carrageenan and the green seaweed produces ulvan. One of the interesting components of seaweed is resident in the external sugar, mainly polysaccharides, in addition there are some minerals, lipids, vitamins and pigments. These MPS change

according to the different seaweed families. Moreover the presence of a sulfate increases their interest. The sulfated polysaccharides extracted from macro algae have demonstrated several pathogenic control properties, such as: antiinfectious, antiviral (against ISA in salmon), anti-bacterial, anti-tumoral, antioxidant and immune-modulation. The above shows that marine sulfate polysaccharides stimulate the immune response and boost the activity of immune cells in order to mitigate the influence of pathologies. Recent studies demonstrate that sulfated polysaccharides have a role in Toll Like Receptor (TLR) activation and can influence the innate immune response. These activate the receptors called Pattern Recognition Receptors (PRR), or TLRs of phagocytic cells. TLRs are transmembrane

proteins that detect invading pathogens by binding to ancestral molecules of microbial origin, called Pathogen-Associated Molecular Patterns (PAMPs). Polysaccharides, then, provide the highest capacity for carrying biological information, as they have a large structural variability. The watersoluble polysaccharides found in green seaweed of the order Ulvales (Ulva and Enteromorpha), have sulfate, rhamnose, xylose and iduronic and glucuronic acids as their main constituents

Macro algal extracts for practical aquaculture use Some marine macro-algae (green, red and brown, for simple classification) have polysaccharides with functional properties which, when added to feed, enhance the health status of farmed and aquatic animals. These can be summarized in five points:

a) Immune modulation As already cited in the previous paragraphs, several polysaccharides compose the complex algal cell wall. After scientific study it was discovered that compounds enhance the immune activity of aquatic animals. Studies on red algae have been show to improve the immunomodulation activities of aquatic animals, increasing the macrophages phagocytosis. It is well known that in some aquatic animals the primitive immune system has difficulty in fighting some diseases. Also carrageenan helps macrophage


34

The use of seaweed extracts can absolutely be used in aquaculture farming systems. One of the main extract products is the MSP that have complex poly-anionic structures with various beneficial biological properties, already used in the aquaculture sector

activity. These polysaccharides are not present in the majority of terrestrial plants. All aquatic animal diseases where the immune system needs a boost or is impaired have been better controlled with the macro-algal polysaccharides sulfonate additions. Green algae (i.e. Ulvans) activate the expression of some cytokines and chemokines that work in innate and adaptive immune response, providing increased immunomodulation properties of the macro-algae additive. This is useful to have a better performing immune system, particularly in young animals and for a better alternative to vaccination. In fact, for general stimulation of the body’s defense mechanism, regular sequential intakes not connected with vaccination are helpful to strengthen

the immune system. Repeated use allows the development of a periodic use of selected sulfonate polysaccharides, this permits the strengthening of the ‘basic’ immune system and the boosting of the state of defense of the innate system. Also the use of polysaccharides upstream or downstream of a prophylactic program prevent pathologies in high density aquaculture farming. Vaccine prophylactic programs can be enhanced with targeted intakes of algal extract, improving the vaccine’s technical and economic performance.

b) Digestive mucin secretion stimulation Several studies and trials have showed that algal polysaccharides (alginate and ulvan) can increase mucin secretion. Intestinal mucins are glycoproteins that cover the luminal surface of the enteric epithelium. Thus they protect the intestinal tract with a gel against the mechanical problem that could occur, such as selfdigestion, bacterial toxins and bacteria, improving aquatic animal gut health. The acetate and butyrate originated from the fermentation of sulfonate polysaccharides also has similar properties. All this must be considered as academic, as responses will be different in each farming situation and every additive used.

c) Digestive health: fat metabolism management Another interesting property of certain macro-algal extracts is their ability to considerably help fat

assimilation; in humans they are used to lower cholesterol levels, due to the high ion-exchange capacity.

d) Prebiotic activities Algal polysaccharides can bypass digestion and serve as a substrate for microbial growth at the lower part of the gut, passing the gastric digestion without being degraded. So polysaccharides can work as prebiotics. Alginates, laminarin and fucans are some of the most interesting polysaccharides present in macro algae, sometimes there are different mix extracts of the different polysaccharides for a more stringent practical effect in farmed animals. In some experiments, algal polysaccharides (laminarin) enhance the development of beneficial bacteria in the gut against those of pathogenic orientation. The above can provide substantial feed cost saving and promote growth, normally obtained with antibiotics. These brown algae can provide a valid alternative to antibiotics, a major problem today.

e) Mycotoxin control: clay and algal polysaccharides interaction The use of macro algal polysaccharides together with modified, finely milled clay, increases the absorptive capacity of clay. This is very useful in mycotoxin binding and can solve the mycotoxin problem in aquaculture feed. Moreover it is possible to use only 1 Kg of clay per tonne of additive against the 10 Kg without algal polysaccharides for a standard mycotoxin treatment.


35

Conclusions The use of seaweed extracts can absolutely be used in aquaculture farming systems. One of the main extract products is the MSP that have complex poly-anionic structures with various beneficial biological properties, already used in the aquaculture sector. In fact, practical use and a number of scientific studies have already highlighted the benefits; particularly the fucoidans, the carrageenan and the ulvans, that control some inflammatory response mechanism and has favorable benefits to aquatic animal immunity. These molecules are already used as agents for the stimulation of the various mechanisms associated with the body defense and, in particular, of the innate immunity mechanisms.

Moreover it must also be considered that the macro-algal extracts are also used in the aquaculture sector for digestive mucin secretion stimulation, fat metabolism management, prebiotic activities, and, when used in association with clay, mycotoxin. control.

It is also possible to use the MPS extract for the fish processing industry for its biocide characteristics as a coadjutant and additive.

More studies on different species of aquatic animals to further refine MSP uses in aquaculture are ongoing. The use of the seaweed extract in aquaculture can boost the production of aquatic animals, while meeting biological legislation and using less antibiotics. References available on request.

More information Gianluigi Negroni E: gigineg@gmail.com

Worldwide jobs For the aquafeed & allied industries Visit the JOB CENTER at aquafeed.com EMPLOYERS: send your jobs to news@aquafeed.com

AFâ„Ś


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Zeroing out the environmental impacts of aquafeed: The Veramaris solution By Ester Santigosa, PhD., DSM Nutritional Products France

Fish oil from wild-caught fish has been the only commercially available source of omega-3 fatty acids until now, driving the aquaculture industry to look for sustainable solutions to meet soaring demand. Without a change, the demand for omega-3 from a global population of ten billion by 2050 will outstrip the world’s natural marine resources. There will not be enough forage fish –sardines, anchovies and sprat – in the ocean, to support aquaculture’s current demand for Omega-3 EPA & DHA fatty acids (Tacon and Metian, 2008; Naylor et al., 2009; Tsikliras et al., 2013). And this will certainly limit the growth of the aquaculture industry. Almost 20% of global wild fish catch is processed into fish oil and fishmeal to feed farmed fish.

Veramaris challenge With the global demand for healthy seafood increasing annually by 3.2% (FAO, 2018), and with consumers becoming increasingly health conscious, many people are looking at including more fish in their diet to get omega-3. Oily fish, especially salmon, are a rich source of the two-essential omega-3 fatty acids, EPA and DHA. To enable aquaculture to provide enough fish for a growing world population, we must close the supply-demand gap for omega-3. That is the challenge Veramaris set to solve, when creating a new technology to provide a highly sustainable and healthy source of EPA

and DHA that does not impact marine life and enables aquaculture to grow. With the help of natural marine microalgae, Veramaris bypasses the traditional aquaculture food chain. EPA and DHA provided by Veramaris are safe, digestible and are well deposited on the fillet: different trials in salmon and trout show consistent results in all the parameters studied, relevant for the aquaculture industry.

Trout trials Trout fed with a diet containing 41.6% protein and 24.7% lipid and including 6% of Veramaris oil as the only source of omega 3 showed no differences in


37 growth parameters when compared to a commercial-like diet with 10% fish oil and 25% fish meal after 12 weeks of experimental feeding (507.9 ± 6.7 versus 506.5 ± 5.7, respectively). Fish fed with diets including Veramaris oil at 3%, 4.5% and 6% were in good health condition as stated by external fish veterinarian.

Salmon trials Juvenile salmon (415g) were fed with a commercial-like diet with 10% fish meal. Digestibility of fatty acids and fatty acid muscle profile were compared to salmon fed diets with 2.5%, 5%, 7.5% and 10% replacement of fish oil by Veramaris oil. Results show that digestibility of DHA is significantly increased in fish fed the three diets with the highest replacement levels (+1.39; +1.28 and +1.42% respectively when compared to control diet; Fig.1). Similarly, EPA+DHA deposition in the fillet increases from 22.40 to 36.90 mg EPA+DHA/g of flesh (Fig. 1) when dietary content increases from 15.2 to 59.8 mg/g feed. This is very important from a product quality point of view, since it has been stated that fish fed low EPA+DHA levels show higher frequency of fillet melanin spots, possibly related to of inflammation (Sissener et al., 2016a). Similarly, salmon (140g) fed for 4 months diets with equal EPA+DHA content (19.54 ± 0.4 mg/g feed) provided by 0, 25, 50 and 100% of Veramaris oil (a maximum of 3% Veramaris oil inclusion) shows a tendency to increase DHA and EPA+DHA ADC with increasing

Figure 1: Apparent digestibility coefficient (up) and omega-3 content in muscle (down) of basal diet (Diet A) and diets with 25, 50, 75 and 100% of fish oil replacement by Veramaris oil (Diets B, C, D and E, respectively). Significant differences between treatments (p-value < 0.05 in one –way ANOVA statistical test) are shown by different letters. NS = non significant differences.

inclusion of Veramaris oil. Literature describes that DHA digestibility can be reduced depending on the dietary lipid source (Karalazos et al., 2011; Caballero et al., 2002: Francis et al., 2007; Hatlen et al., 2012). Our results show that EPA and DHA provided by Veramaris oil are highly digestible as previously stated by Miller et al. (2007), using Schizochytrium oil. Omega-3 fatty acid profile in the muscle reflects the dietary fatty acid profile, with no significant differences among treatments (Fig. 2) as widely described previously in the literature.

Results Our data show that, when provided by Veramaris microalgae oil, EPA and DHA are very well digested: apparent digestibility coefficients above 98%. Similarly, when levels of EPA+DHA in the diet are maintained whatever the omega-3 source is, the muscle fatty acid profile is unaffected. As described previously in literature, muscle fatty acid profile reflects the dietary profile. Moreover, when EPA+DHA dietary content increases - even at high levels up to 59.8 mg EPA+DHA/ g feed -,


38 Xu et al, 2014 showed that DHA/EPA ratio in the diet may influence certain innate immune responses and stress tolerance in seabass. This ratio also affects also lipid metabolism, antioxidant capacity and hematological characteristics in black seabream (Jin et al., 2017).

Figure 2: Apparent digestibility coefficient (up) and omega-3 content in muscle (down) of basal diet (Diet A) and diets with 25, 50, 75 and 100% of fish oil replacement by Veramaris oil (Diets F, G and H, respectively). Significant differences between treatments (p-value < 0.05 in one–way ANOVA statistical test) are shown by different letters. NS = non significant differences.

DHA is deposited in a linear way and EPA is also increased in the fillets. These results support the fact that our omega-3 from microalgae is an innovative, premium alternative to fish-based feed ingredients. The Veramaris omega-3 oil will alleviate the pressure on marine habitats and conserve the biodiversity of our oceans. We encourage industry leaders to adopt sustainable feed production as a worldwide standard and become stewards of the ocean. By doing this, they will improve the sustainability and health profile of

seafood since natural marine algal oil is particularly rich in omega-3 EPA.

Demonstration program Veramaris is now developing a research program aiming to demonstrate the benefits of both EPA and DHA to the animals. Preliminary work in salmon (Sissenser et al.,2016b) has suggested that it is important to distinguish between requirements for DHA and requirements for EPA. In this sense,

We face a global challenge of how to feed a growing population without exhausting precious natural resources. Veramaris’ breakthrough solution will enable the aquaculture industry to decrease its reliance on wild caught feeder fish and become a net fish producer – the holy grail of the aquaculture industry. As the aquaculture industry continues to grow, consumers must understand that not all farmed fish will deliver the same benefits. If we want to ensure that the aquaculture industry becomes more sustainable and can continue to provide the health benefits to the consumers; then, we need to educate them on the importance of EPA and DHA as health promoters.

References Caballero M.J., Obach A., Rosenlund G., Montero D., Gisvold M., Izquierdo M.S. (2002) Impact of different dietary lipid sources on growth, lipid digestibility, tissue fatty acid composition and histology of rainbow trout, Oncorhynchus mykiss. Aquaculture 214:253–71. FAO. 2018. The State of World Fisheries and Aquaculture 2018 Meeting the sustainable development goals. Rome.


39 Francis D.S., Turchini G.M., Jones P.L., De Silva S.S. (2007) Growth performance, feed efficiency and fatty acid composition of juvenile Murray cod, Maccullochella peelii peelii, fed graded levels of canola and linseed oil. Aquaculture Nutrition 13: 335–350. Hatlen et al., 2012. Hatlen B., Berge G.M, Odom J.M., Mundheim H., Ruyter B. Growth performance, feed utilisation and fatty acid deposition in Atlantic salmon, Salmo salar L., fed graded levels of high-lipid/high-EPA Yarrowia lipolytica biomass. Aquaculture 364–365, 39–47. M. Jin, O. Monroig, Y. Lu, Y. Yuan, Y. Li, L. Ding, D.R. Tocher, Q. Zhou (2017). Dietary DHA/EPA ratio affected tissue fatty acid profiles, antioxidant capacity, hematological characteristics and expression of lipid-related genes but not growth in juvenile black seabream (Acanthopagrus schlegelii) PLoS One, 12 (4) (2017), Article e0176216 Karalazos V., Bendiksen E.A., Bell J.G. (2011) Interactive effects of dietary protein/lipid level and oil source on growth, feed utilisation and nutrient and fatty acid digestibility of Atlantic salmon. Aquaculture 311, Issues 1–4, 3: 193–200. Miller M.R., Nichols P.D., Carter C.G. (2007) Replacement of fish oil with Thraustochytrid Schisochytrium sp. oil in Atlantic salmon parr (Salmo salar L.) diets. Comparative Biochemistry and Physiology – Part A 148: 382–392. Naylor RL, Hardy RW, Bureau DP, et al. (2009) Feeding aquaculture in an era of finite resources. Proc Natl Acad Sci U S A 106, 15103–15110.

Sissener N, Waagbø R, Rosenlund G, Tvenning L, Susort S, Lea TB, Oaland Ø, Chen L, Breck O. (2016a). Reduced n-3 long chain fatty acid levels in feed for Atlantic salmon (Salmo salar L.) do not reduce growth, robustness or product quality through an entire full scale commercial production cycle in seawater. Aquaculture 464. 236-245. Sissener, N.H.; Torstensen, B.E.; Stubhaug, I.; Rosenlund, G. (2016b) Long-term feeding of Atlantic salmon in seawater with low dietary longchain n-3 fatty acids affects tissue status of brain, retina and red blood cells. Br. J. Nutr. 115, 1919–1929.

Veramaris, based on natural marine microalgge, provides highly sustainable and healthy source of DHA and EPA that is safe, digestible and well deposited bon fish fillet

Tacon AGJ & Metian M (2008) Global overview on the use of fish meal and fish oil in industrially compounded aquafeeds: Trends and future prospects. Aquaculture 285, 146–158

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Tsikliras AC, Dinouli A & Tsalkou E (2013) Exploitation trends of the Mediterranean and Black Sea fisheries. Acta Adriatica 54,273–281.

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Xu, H.; Wang, J.; Mai, K.; Xu, W.; Zhang, W.; Zhang, Y.; Ai, Q. (2014). Dietary docosahexaenoic acid to eicosapentaenoic acid (DHA/EPA) ratio influenced growth performance, immune response, stress resistanceand tissue fatty acid composition of juvenile Japanese seabass, Lateolabrax japonicus (Cuvier). Aquac. Res. 2014, 3, 741–757.

Learn more about this work at Aquafeed Horizons, June 12 2019, Cologne, Germany. More information: http://feedconferences.com

More information

Dr. Ester Santigosa Senior Scientist Aquaculture DSM Nutritional Products France Florida, USA E: ester.santigosa@dsm.com


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Trends and Developments Changing your feed? You might want to change your fish also

Photo: Citron / CC-BY-SA-3.0

By C. Greg Lutz, PhD.

Gains resulting from the processes of domestication and selection have leveled out for most terrestrial animal industries over the past few decades, and current practices reflect a combination of adapting the animals to their environments and vice versa. The potential for significant genetic improvement still remains relatively

untapped for many aquaculture species but as has been the case for poultry, pork and beef production, approaches that combine disciplines such as physiology, genetics and nutrition will probably result in the most important advances in the coming years. One example worth noting involves

recent research in France, focused on utilizing fish-free diets for rainbow trout. In 2015, Viviana Lazzarotto et al. published an article in PLoS ONE examining the reproductive consequences of feeding an entirely plantbased diet to rainbow trout (Oncorhynchus mykiss). When Lazzarotto et al. began their work,


42 that n-3 LC-PUFA requirements were met.

... after only three generations of selection, selected fish fed the plant-based diet reached a similar final body weight as the control fish fed the marine resources-based diet, along with an improved survival at early stages. Similar results have been reported for other salmonids. The basic message here is: if an industry intends to change its feed, itâ&#x20AC;&#x2122;s probably wise to also consider changing the fish. Weâ&#x20AC;&#x2122;ll undoubtedly see more of this interdisciplinary approach over the next decade.

previous studies with rainbow trout had shown that the complete replacement of fishmeal with plant protein led to slower growth. This was probably linked to a combination of decreased feed intake, poorer digestibility of carbohydrates and the presence of anti-nutritional factors. However, several studies carried out in salmonids had shown that it was possible to totally replace fish oil with vegetable oils without affecting growth or feed efficiency, provided

Lazzarotto et al. wanted to determine whether rainbow trout reared on a plant-based diet from first feeding to reproduction were capable of synthesizing sufficient amounts of n-6 and n-3 LC-PUFAs to survive and reproduce. Trans-generational effects on offspring survival were also studied to assess whether female trout reared on a plant-based diet could produce viable ova and offspring. The experiment was conducted by rearing female fish using either a commercial (C) diet in which 45% of the fishmeal and 50% of the fish oil were replaced by plant ingredients, or an experimental plant-based diet (V), completely free of fishmeal and fish oil. After their first (year-2) spawning season, the researchers continued raising the fish up to their next spawning (at 3 years old). At each spawning approximately 400 ova per female were fertilized with a pool of sperm collected from males fed a commercial diet. Survival rates of progeny were measured at the eyed stage, at hatching, and at the swim-up fry stage. At first spawning the body weights of females fed the V-diet were higher (+18%) than those of females fed the C-diet, indicating that limitation of food supplied to the C fed group may have been too restrictive. No such difference was found at the year-3 spawning. At first spawning, the absolute fecundity of the V-fed females was higher (+17%) than that of the C-fed females. However, at the year-3 spawning no such difference was detected. Average ovum weight

was significantly lower in V-fed fish at both spawnings, as was the gonadosomatic index. At the first spawning, survival rates for progeny from V-fed females were lower than from broodstock fed the C-diet, but at the year-3 spawning no significant differences were detected.

Addressing both fishmeal and fish oil Wide variability was observed in the performance of female trout fed the vegetarian diet, so a separate study was designed by Therese Callet (et al.). Their findings were published in 2017, also in PLoS ONE. Various studies had demonstrated the existence of genetic variability in growth of rainbow trout populations when fed increasing proportions of plant products, with some genotypes able to accept and use these diets more readily than others. Some studies had also demonstrated interactions between diets and genotypes when comparing growth, feed intake and feed efficiency. Most research in this area had only focused on fishmeal replacement, not fish oil. Callet et al. decided to address the question of replacement of both fishmeal and fish oil, and selection was imposed for the ability to survive and grow with a plant-based diet completely devoid of marine ingredients, from the very first feeding. The base population for Callet et al.â&#x20AC;&#x2122;s study was a domesticated strain fed over generations with a commercial diet containing a mix of marine and


43 plant ingredients. From this line, a new line was selected over three generations for its ability to survive and grow when fed a plant-based diet with no fish oil or fishmeal. Fish obtained from these crosses were fed from the first-feeding stage with a strictly plant-based diet. Marine products were replaced by a blend of vegetable meals (wheat, fava bean, corn, soybean, lupin and peas) and vegetable oils (rapeseed, linseed and palm oils). Plant-based diets were supplemented with amino acids to meet known rainbow trout essential amino acid requirements, and with a mineral and vitamin mix. Surviving fish were graded by size several times between the second and fourteenth months after first feeding. The final proportions of selected fish ranged from 3.9% (first generation) to 4.9% (third generation). After the last sorting (14 months), selected fish were fed a commercial diet to ensure normal reproduction and prevent any selection on fish reproductive capacity with the V diet since Lazzarotto et al. had shown that rainbow trout females fed all their life with a 100% plantbased diet had very heterogeneous reproductive performances. The shift to the commercial diet was made after the last sorting, so this diet should not have influenced selection results. A 7-month feeding trial was performed on fish produced from the third generation of selection. From first feeding (41 dpf) to 197 dpf, within each line three batches were fed with a 100% plant-based diet while the three other batches were fed with a marine resources-based

diet. At each sampling date, there was an effect on growth from both the diet and the selection history. While the plant-based diet diminished fish body weight, at least partially due to lowered feed intake, selection imposed on prior generations increased it. There were no significant differences between the mean body weight of selected fish fed the plantbased diet and control fish fed the marine resources-based diet from 78 dpf until the end of the trial. Still, the majority of the nutritional traits studied were highly impaired by the plant-based diet, in both lines. Increased fish performance was attributed to a higher feed intake, irrespective of the diet. This result highlights the fact that acceptance of plant-based diets is a major step impeding the replacement of fish oil and fishmeal, but it can be significantly improved by selection for appetite. The most significant finding of the Callet et al. study was that after only three generations of selection, selected fish fed the plant-based diet reached a similar final body weight as the control fish fed the marine resources-based diet, along with an improved survival at early stages. Similar results have been reported for other salmonids. The basic message here is: if an industry intends to change its feed, it’s probably wise to also consider changing the fish. We’ll undoubtedly see more of this interdisciplinary approach over the next decade. For further details on the studies mentioned here, see: Lazzarotto V, Corraze G, Leprevost A, Quillet E, Dupont-Nivet M, Médale F

(2015) Three-Year Breeding Cycle of Rainbow Trout (Oncorhynchus mykiss) Fed a Plant-Based Diet, Totally Free of Marine Resources: Consequences for Reproduction, Fatty Acid Composition and Progeny Survival. PLoS ONE 10(2): e0117609. doi:10.1371/ journal.pone.0117609

Callet T, Médale F, Larroquet L, Surget A, Aguirre P, Kerneis T, Labbe L, Quillet E, Geurden I, Skiba-Cassy S, Dupont-Nivet M (2017) Successful selection of rainbow trout (Oncorhynchus mykiss) on their ability to grow with a diet completely devoid of fishmeal and fish oil, and correlated changes in nutritional traits. PLoS ONE 12(10): e0186705.

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About the author

Dr. Greg Lutz is a Professor with Louisiana State University Agricultural Center. He is also an author and consultant, and serves as the Editor in Chief of Aquaculture Magazine. E: lutzaqua@att.net


HE 6TH GLOBAL EED AND FOOD CONG RESS 2019

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NGKOK, THAILAND. 11—13 MARCH

THE FUTURE OF FEED & FOOD ARE WE READY? Date & Venue

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The 6th Global Feed & Food Congress (GFFC), organized by the International Feed Industry Federation (IFIF) with technical support provided by the Food and Agriculture Organization of the United Nations (FAO) and in collaboration with VIV worldwide will be held at the exclusive Shangri-La Hotel in Bangkok, Thailand, on 11-13 March 2019.

The 6th GFFC theme ‘The future of Feed & Food – are we ready?’ links to the global challenge to provide safe, affordable, nutritious and sustainable animal protein sources through innovative solutions to feed 9 billion people by 2050 and reflects our shared vision to achieve this for a growing world population now and for the future.

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The 6th GFFC will look ahead at key topics for the feed & food chain, including:

The 6th GFFC is expected to attract executive level delegates from Asia, Europe, Africa and the Americas. Join us and you will: — Experience exceptional speakers who will share their insights on the future of feed and food. — Network and discuss strategy with business leaders, senior government officials, experts and policy makers from the feed & food value chain. — Engage with leading animal nutrition and food companies, food chain partners, international organisations, national authorities and international civil society.

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45

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Interesting nutrition studies in the Chinese literature By Albert G. Tacon, Ph.D., Technical Editor, Aquafeed.com

Aloha. As we all know, China is the giant on the block with regard to aquaculture production. It follows therefore that their research community is also very active, with the majority of research papers being published in Chinese journals for the benefit of the Chinese aquaculture community. Since most of these publications also have english abstracts, I thought it would be useful to share these with our readers. Sometimes the abstracts are not written in the best English possible, but I think that they are worth sharing. Here is a small sample of some of their publications that I think will be of interest. I hope that this will be a regular feature of this column depending on your interest! Effect of small peptide supplementation at different protein levels on growth performance, digestive enzymes activities, serum biochemical indices and antioxidant abilities of grouper (Epinephelus akaara). Zhao, Shuyan; Lin, Heizhao; Huang, Zhong; Zhou, Chuanpeng; Wang, Jun; et al. (2016). South China Fisheries Science, 12(3). Six diet groups containing protein (44%, 48%, 52%) and small peptide (0.05% and 0.1%) were fed to groupers (Epinephelus akaara) to assess the effect on growth perfor-

mance, digestive enzymes activities, serum biochemical and antioxidant abilities. The diets were named as D1 (44%, 0.05%), D2 (44%, 0.1%), D3 (48%, 0.05%), D4 (48%, 0.1%), D5 (52%, 0.05%) and D6 (52%, 0.1%). Each diet was fed to triplicate groups of fish with 15 fish [initial weight (42. 16 + or - 0. 23) g] which were randomly sorted into each rearing cubic fiberglass tanks and the experimental last for eight weeks. The results show that there was no effect on weight gain (WG), specific growth rate (SGR),muscle moisture, crude

Dr. Albert Tacon is the Technical Editor of Aquafeed.com and an independent aquaculture feed consultant. E: agjtacon@aquahana.com

protein and ash contents. As protein level and small peptide concentration increased, the crude fats of grouper muscle, triglyceride (TG) and cholesterol (CHO) decreased (P 0.05).The activity of pepsin was the highest in Group D2,and the activities of amylase, trypsin and lipase were the highest in Group D1.There were no significant difference among different groups (P 0.05). The activities of malondialdehyde (MDA) and peroxidase (POD) were not significantly different among different groups (P 0.05). The result shows that


46 supplementation of small peptide at different protein levels can reduce protein requirement, promote fat metabolism and increase digestive enzymes activities.

Effect of dietary Rhodotorula mucilaginosa on growth performance, digestive and immune enzymes activities of juvenile Nile tilapia (Oreochromis niloticus). Yang, Keng; Lin, Heizhao; Xia, Dongmei; Zhou, Chuanpeng; Yang, Yingying; et al. (2016). South China Fisheries Science, 12(6). A growth trial was conducted to investigate the effect of dietary Rhodotorula mucilaginosa on the growth performance, digestive and immune enzymes activities of juvenile Nile tilapia (Oreochromis niloticus). Six diets were formulated by adding different levels of R. mucilaginosa to basal diet (control) : 1 g/kg, 2 g/kg, 3 g/kg, 4g/kg and 5 g/kg, respectively. The results show that compared with the control, the FCR of the fish in 2 g/ kg and 4 g/kg groups decreased significantly (P 0.05). The body crude protein of the fish in 5 g/kg group was significantly lower than those in 1 g/ kg and 2 g/kg groups (P 0.05). The body crude ash of the fish in 1g/kg, 3 g/kg, 5 g/kg groups were significantly lower than that in control group (P 0.05). Compared with the control, the pepsin activity of the fish in 1g/kg group increased significantly (P 0.05), but the pepsin activities of the fish in 3 g/kg, 4 g/kg and 5 g/kg groups decreased significantly (P 0.05). The amylase activities in stomach, liver, foregut and midgut in 2g/kg group

were higher than those in control group significantly (P 0. 05). Compared with the control, the hepatic lysozyme and nitric oxide synthase activities of the fish in 2 g/kg and 3 g/ kg groups increased significantly (P 0.05). In conclusion, the optimal dietary R. mucilaginosa level in juvenile tilapia feed could be 2 ~ 3 g/ kg.

Effect of replacing fish meal with soybean protein concentrate on growth, body composition, serum biochemical indices, and liver histology of juvenile large yellow croaker (Larimichthys crocea). Feng, Jian; Wang, Ping; He, Jiaojiao; Lou, Yudong; Dang, Hui. (2017). Zhongguo Shui Chan ke xue Beijing, 24(2):268. A 56-d feeding experiment was conducted to investigate the effects of replacing fish meal in the diet with soy protein concentrate (SPC) on growth, body composition, serum biochemical indices, and liver histology of juvenile large yellow croaker, Larimichthys crocea (initial weight 10.50 g). The basal diet contained 40% fish meal, and five iso-nitrogenous (crude protein 45%) and isolipidic (crude lipid 10%) diets were formulated by replacing 0%, 25%, 50%, 75%, and 100% of the fish meal with SPC and were designated the FM, R25, R50, R75, and R100 groups, respectively. All diets were supplemented with the appropriate amounts of crystalline lysine and methionine, based on lysine and methionine contents in the control group. Each diet was assigned randomly to triplicate groups of 60 individuals/cage. All larvae were

stocked into experimental cages (1.5 m x 1.5 m x 2 m) at Xihugang, Xiangshan County, Ningbo city, Zhejiang Province, China.

Allometric growth and ontogenetic changes in nucleic acids and digestive enzymes during the early life stage in fish species: A review. Zhang, Yunlong; Zhang, Hailong; Wang, Lingyu; Gu, Beiyi; Fan, Qixue. (2017). Zhongguo Shui Chan ke xue; Beijing, 24(3):648. The early life stage is a critical period in fish species since complicated physiological and morphological changes and often massive mortalities occur at this stage. Studies of the growth patterns and physiological characteristics during the early life stage often uncover lethal factors in this stage, and the information gained can lead to improvements in survival rates during larval production. Allometric growth can be used as an indicator for larval production, and fish experience a change in shape in relation to increases in their ability to perform vital biological functions needed for survival during their early life stage. The RNA/DNA ratio is a sensitive indicator of growth rates which can be used to evaluate growth potential and nutritional condition, as well as to determine critical periods in the larval stage. Specifically, studies on the development of the digestive tract and digestive capability of the organism can be used as an indicator of nutritional status at an early life stage, thereby providing information useful either for improving feeding protocols in larviculture, suggesting


47 more suitable food items, or designing feasible larval rearing procedures.

Effect of dietary protein level on growth performance, body composition, and digestive enzyme activities in green mud crab (Scylla paramamosain) juveniles. Dong, Lanfang; Tong, Tong; Zhang, Qin; Xu, Mingzhu; Su, Qiong; et al. (2017). Zhongguo Shui Chan ke xue; Beijing, 24(3):524. The green mud crab Scylla paramamosain has been widely cultured in coastal areas of southern China for many years, with production reaching approximately 14.1 tons in 2015.The successful farming of mud crabs presently depends on conventional diets, comprising mainly trash fish, mollusk meat and animal viscera, but the demand for these foods by the crab industry is difficult to satisfy. Compared with the traditional feeds, compound feeds offer better-balanced nutrition and have better storage qualities. Studies of the nutritional requirements of cultured mud crab are important to developing costeffective, environmentally friendly and nutritionally balanced artificial diets. Dietary protein, a major macronutrient in aquatic animal diets, provides the essential and nonessential amino acids needed to synthesize body protein and energy. As protein represents the most expensive component in feeds, it is important to determine the optimal level for a cultured species. This study investigated the effects of dietary protein level on growth performance, body composition, and digestive enzyme activities in mud crab (S. paramamosain) juveniles. Six

isoenergetic diets were formulated with varying protein content (i.e., 31.45%, 36.37%, 41.55%, 46.13%, 51.72% and 56.86%) and fed to juvenile mud crabs for three weeks. Triplicate groups of 80 juvenile crabs, with an average body weight of 11.86 ~c 0.11 mg, were randomly assigned to each of the six diets. Each crab was raised in a bucket (20 cm diameter Ă&#x2014; 25 cm height) provided, with a 5 cm layer of sand. The results showed that the dietary protein level did not influence crab survival rate in any of the treatments (P<0.05). The weight gain ratio and specific growth rate of the juvenile crabs significantly increased as dietary protein was increased from 31.45% to 51.72% (P<0.05),and then the indicators showed a slight decrease with a further increase to 56.86% protein; the gains were highest for crabs fed 51.72% protein.

Effects of dietary fat and protein nutrition on fat metabolic enzymes and growth-associated gene expression of Atlantic salmon (Salmo salar) in the recirculating aquaculture system. Ma, Jun; Liu, Yang; Li, Yong; Zhang, Jing; Zhao, Ningning. (2017). Zhongguo Shui Chan ke xue; Beijing, 24(4):669. Salmo salar is one of the most important aquaculture species in the world, cultured in a cage culture system, which was introduced to China in 2010 and in recirculating aquaculture system (RAS). In order to investigate the effects of dietary fat and protein levels on fat metabolic enzymes and growth associated gene expression of S.salar in RAS, a 3x 2

(three fat levels: 18%, 21%, and 24%; two protein levels: 38% and 48%,represented by F18, F21, F24, P38, and P48, respectively) random two-factor animal trial was conducted for 56 days. The trial included six treatments with triplicate groups of 40 trial fishes (650.00+45.50 g) Results showed that: (1) The fat requirement of S.salar was lower in RAS than in cage culture system, and the requirement of protein was equal between the two systems. The weight gain rate (WGR) was significantly higher in P48 F21, by 22.23%125.86%, than that in other groups (P<0.05), while the feed conversion rate (FCR) was significantly lower in P48 F24, by 16.24%-30.00% than that in other groups (P<0.05). (2) High fat level significantly increased hepatosomatic index (HSI) (P<0.05), while high protein level decreased condition factor (CF) very significantly (P<0.01). The HSI was significantly higher in P48 F24 group than in other groups, by 10.92%-28.16% (P<0.05), while the CF was significantly lower than that in other groups, by 10.24%-12.31%. The study proposed the primary nutrition project that can control the shape of S.salar to 600-900 g by weight. (3) High and medium fat levels significantly (P<0.05) increased the activity of lipoprotein lipase (LPL), hepatic lipase (HL), and total esterase, while high protein level significantly (P<0.05) increased the activity of LPL and total esterase.

A comparative study on growth and gonadal development of Litopenaeus vannamei broodstock fed different formulated feeds. Zhang, Yuling; Luo,


48 Kun; Kong, Jie; Liang, Mengqing; Luan, Sheng; et al. (2017). Zhongguo Shui Chan ke xue; Beijing, 24(2):306.

Effects of biofloc technology on growth performance and disease resistance of gibel carp, Carassius auratus gibelio. Zhang, Mingming; Xu, Chen; Zhao, Zhigang; Wang, Zisheng; Qiu, Ming; et al. (2017). Zhongguo Shui Chan ke xue; Beijing, 24(3):533.

water exchange, environmentally friendly aquaculture system that is now widely used since it is proven to enhance the growth and immunity of farmed fish. Gibel carp Carassius auratus gibelio (also known as Prussian carp) is a major aquaculture species in China, especially around Yancheng in Jiangsu Province, and it is also grown in mudflats. Developing an effective aquaculture system for gibel carp is also helpful for exploring mudflats usage in fish aquaculture. However, sudden outbreaks of disease have become a great and common threat to carp-farming operations, as this causes huge economic losses. Since 2009, an epizootic with severe mortality rates caused by cyprinid herpes virus 2 (CyHV-2) has emerged among cultured gibel carp in China, especially in the Northem part of Jiangsu Province. The disease develops fast, spreads widely to almost all main areas of gibel carp, and mortalities may reach 90%-100%. Thus, disease becomes a limiting factor for aquaculture, yet effective methods to prevent or control CyHV-2 are lacking. In order to discuss the practical usage of BFT in mudflat fish culture in terms of disease prevention, this study subjected the representative species (gibel carp) to the disease, using fish cultured in two different systems: in a BFT system without any water exchange, and in a common recirculating system with 1/4-1/3 water exchange daily (as control), to analyze the effect of BFT on growth performance and disease resistance in this carp.

Biofloc technology (BFT) is a zero-

Changes in histology and protein

This experiment was conducted to evaluate different artificially formulated feeds on promoting gonadal development in Litopenaeus vannamei broodstock during the nutrient enrichment stage. Three kinds of formulated feeds, numbered diets no.1 (without krill meal), no.2 (10% krill meal), and no.3 (20% krill meal) were used as the experimental groups, with two replicates, and natural Nereis polychaete worm bait was used as the control group. The purpose of the experiment was to determine the appropriate formulated feed to enhance gonadal development in L.vannamei broodstock. The four treatment groups were fed the four different feeds for 60 d, and wholebody, hepatopancreatic, and gonad weights were determined. In addition, the fatty acid profiles of the four feeds and those of the hepatopancreas and gonads were analyzed in the four treatment groups. Fatty acid profiles of feed are important for gonadal development and reproduction in L.vannamei brood stock, as they can affect the hepatopancreatic and gonadal fatty acid profiles, as well as gonadal development.

content in claw closer muscle of Chinese mitten crab, Eriocheir sinensis during the molt cycle. Tian, Zhihuan; Jiao, Chuanzhen; Wu, Xugan; Cheng, Yongxu. (2017). Zhongguo Shui Chan ke xue; Beijing Vol. 24, Iss. 5, (2017): 1072. To characterize the changes in the protein content and tissue structure of the claw closer muscle of Eriocheir sinensis during the molt cycle, muscles harvested at various stages during the molt cycle were studied using paraffin section histology, electron microscopy, and biochemical methods. The results showed that the histology of the claw closer muscle changed noticeably during the molt cycle. The fibers and myofibrils were fully expanded in the inter-molt stage, and the cross-sectional area resembled an irregular circle or polygon with many nuclei under the sarcolemma. The sarcomeres were generally in register in this stage. During the later pre-molt stage, the cross-sectional area of fibers showed a wide size distribution owing to pro-ecdysial muscle atrophy, and many eroded areas appeared within the myofibrils. In the post-molt stage, the muscle fibers were compacting and shrinking.

Suitable dietary starch source and supplementation level for largemouth bass (Micropterus salmoides). Liu, Zike; Chen, Naisong; Wang, Mengle; Lian, Xueyuan; Yan, Chunwei; et al. (2017). Zhongguo Shui Chan ke xue; Beijing, 24(2): 317. Eight iso-nitrogenous and isoenergetic (crude protein 48% and gross energy 18.5 MJ/kg) diets (L5,


49 L10, Z5, Z10, X5, X10, M5, and M10) were formulated with 5% and 10% waxy corn starch (L), high-amylose corn starch (Z), wheat starch (X), and cassava starch (M) to determine a suitable starch source and supplementation level for a largemouth bass, Micropterus salmoides, diet. Fish (initial body weight 23.46 g ~c 0.19 g) were fed to apparent satiation twice daily for 45 d. The results showed that the supplemented dietary starch level and starch source had significant effects on growth, feed utilization, body composition, and non-specific immunological indices of largemouth bass. Significant decreases in specific growth rate and feed intake were observed with increasing content of the same starch from 5% to 10%. However, feed and protein efficiency increased significantly in the L10, X10, and M10 groups, compared with those in the L5,X5 and M5 groups, whereas no differences were detected in these parameters between the Z5 and Z10 groups. Hepatosomatic index, viscerosomatic index, and liver glycogen concentration increased significantly in the L10,X10,and M10 groups, compared with those in the L5,X5,and M5 groups, but no

difference was found in the viscerasomatic index between the Z5 and Z10 groups Whole-body and liver lipid contents, as well as liver protein content decreased significantly when any of the supplemented starches were increased from 5% to 10% in the diets. Blood erythrocyte count and hematocrit in the L10 group, serum triglyceride content in the Z10 group, and hematocrit and serum triglyceride content in the X10 group decreased significantly, compared with those in the L5,Z5,and X5 groups. Serum 50% hemolytic complement (CH50) activity was significantly lower in the Z10 group than that in the Z5 group.

Intraspecific variability in growth and fatty acid composition among five strains of lsochrysis galbana. Cong, Chao; Hu, Dongxue; Sun, Chunxiao; Qiao, Hongjin; Wang, Jiying; et al. (2017). Zhongguo Shui Chan ke xue; Beijing, 24(4): 774. Isochrysis galbana is an excellent unicellular bait for the larvae of many aquaculture animals. Thus, screening for I.galbana strains with rapid growth rate and high content of docosahex-

aenoic acid (DHA) and eicosapentaenoic acid (EPA) is very important for the breeding of aquaculture animals. However, most of the I. galbana strains in China have been derived from a common ancestor called I. galbana OA-3011, and different strains of I. galbana have been developed after a long duration of geographical isolation. Whether the traits of these strains have changed or not remains unknown. In order to evaluate the status of the phenotypic traits of different geographic isolates for future selective breeding, comparative analysis of five strains of I. galbana (FACHB-861, OA3011-QD, OA3011-LK, OA3011-AY, and OA3011LZ) with regard to their taxonomy, growth, and fatty acid composition was conducted. Different strains were cultured under identical conditions for six days until they reached a stationary phase. Genomic DNA was extracted for polymerase chain reaction sequencing of the 18S rDNA gene. The OD660 was recorded daily for plotting a growth curve. When the cultures reached stationary phase, they were harvested by high-speed centrifugation for biomass determination and fatty acid analysis by using

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50 gas chromatography. Phylogenetic analysis of the 18S rDNA gene suggested that FACHB-861 was distantly related with the OA-3011 strains. OA3011-LK, OA3011-AY, and OA3011-LZ had identical 18S rDNA sequences, but the 18S rDNA sequence of OA3011-QD differed from that of the other OA-3011 strains at one position, resulting in a single base substitution. The growth curve showed that the five strains reached stationary phase at different times.

Effects of dietary isoleucine levels on activities of digestive enzymes and immune index of Trachinotus ovatus. Huang, Zhong; Zhou, Chuanpeng; Lin, Heizhao; Tan, Xiaohong; Peng, Jingshu; et al. (2017). South China Fisheries Science, 13(1).

We investigated the effects of dietary isoleucine on the digestive enzymes and immune index of juvenile golden pompano, Trachinotus ovatus. Six diets with different concentrations of L-isoleucine (13. 2 g/kg, 15. 7 g/kg,18. 2 g/kg, 20. 7g/kg, 23. 2 g/kg and 25. 7 g/kg dry diet) were formulated to contain 430 g/kg crude protein with fish meal, soybean meal, peanutmeal and pre-coated crystalline amino acids. Each diet was randomly assigned to triplicate treatments of 20 fish [initial body weight was (6. 36 + or - 0. 0 g) for eight weeks. The intestinal amylase and pepsase activities, serum T-AOC and LZM activities, Ig M concentrations increased with increasing isoleucine concentration up to 18. 2 g/kg, whereas decreased in the groups exceeding 20. 7 g/kg isoleucine significantly. Those

parameters of the fish fed with diets of 15. 7 g/kg and 23. 2 g/kg isoleucine were significantly higher than those fed with diets of 13. 2 g/kg and 25. 7 g/kg isoleucine, while significantly lower than groups containing 18. 2 g/ kg and20. 7 g/kg isoleucine. There is an opposite change trend in serum urea and MDA concentration, i. e., significantly lower in 18. 2g/kg and 20. 7 g/kg isoleucine groups than those in the other groups. The SOD activity rate increased with increasing isoleucine concentration up to 18. 2 g/ kg, and there was no significant difference when the dietary isoleucine exceeded 18. 2 g/kg. So diet with 18. 2 ~ 20. 7 g/kg isoleucine can improve the digestive performance and immune function of T. ovatus. AFâ&#x201E;Ś


51

FEED MANAGEMENT

The importance of precise biomass estimation in the feeding of farmed shrimp By Jason Enlow, XpertSea

When done right, shrimp farming can be a profitable business and a sustainable alternative to open sea fishing, but shrimp can be tricky. In the folk tale The Wolf and the Shrimps, some clever shrimp trick a hungry wolf into not eating them. Like the wolf, farmers can be misled by their shrimp, at least in terms of knowing how many there are in a pond. This is important because the estimated number of shrimp will provide the basis for managing and feeding them. Equally critical is being able to determine the right feed conversion ratio (FCR). While check trays and scales can help, many farmers are looking past traditional manual methods to keep their overhead down and optimize their profits. These farmers are embracing recent technological advances in gathering and analyzing data, so they can effectively track metrics such as weight distribution, average weights and weekly growth rates. “In every commercial aquaculture situation, knowledge of actual animal growth rates, feed intake and growth

efficiencies in different environmental conditions, as well as means for estimating and predicting growth, is essential for the viability of the enterprise." 1

The case study: Suyog Aqua Farms—aquaculture shrimp farming Located in Maharashtra, India, Suyog Aqua Farms opened in 2005. Mr. Suyog Powle managed the farm until

2017. During his tenure, the farm annually produced 850 metric tons of L. vannamei shrimp and occupied over 80 hectares of land. The challenge on Suyog’s farm, and on many aquaculture farms, is being able to achieve an optimal FCR (feed conversion ratio). These assessments are sometimes unreliable because they depend on physical observations and are further limited by the technician’s experience level. It is not the best combination when trying to gather data about your ponds.


52 Suyog recalls when one of his ponds reached an FCR of 2.5—a high number when you consider that the general industry rule for L. vannamei shrimp is 1.2 or lower. High FCR is a problem many farmers face when technicians cannot accurately assess feed consumption and thus, have trouble properly comparing it with growth rates. “We thought the shrimp were eating more because the check tray never had any leftover feed. The technician continued to increase the amount of daily feed given but didn’t realize the shrimp were not gaining any weight,” said Suyog Powle.

A check tray is nothing better than a guess To assess feed consumption, technicians at Suyog Aqua Farm use a check tray, a method that Suyog is not impressed with. “A check tray is nothing better than a guess. You really have little way of knowing if what was eaten from the tray is an accurate representation of what’s happening in the pond. Compounding this problem is infrequent manual assessments of the shrimp weight. So, farmers lack critical information about pond biomass, which is required to optimize feeding programs,” said Suyog Powle.

To determine the biomass, the average weight of the shrimp must be calculated and multiplied by the population. On Suyog’s farm, technicians take a sample of shrimp from the pond every 10 days to assess growth using a weighing scale.

Counting shrimp in the XperCount smart bucket, which uses optics, computer vision and machine learning to count, size, weigh and image aquatic organisms, at the press of a button. Photo: XpertSea.

According to Suyog, this information was sporadic and inaccurate. But according to the FAO, weighing might not be a farmer’s best bet when trying to figure out growth rate.

Monitoring shrimp weight is not enough The problem goes beyond the inaccuracies associated with infrequent and manual assessments of shrimp weight. According to section 8 of Feed and Feeding of Fish and Shrimp, a manual published by the FAO (Food and Agriculture Organization of the United Nations): “If accurate length/weight relationships for the species have been predetermined under the environmental conditions being used, length measurements are a more accurate means of monitoring growth rate.” 2 Shrimp can hold uneven quantities of

water in their carapace, which means monitoring weight is not enough to assess their growth with enough accuracy. It is an unfortunate reality, but without the benefit of accurate information, farmers are left with little choice but to base critical decisions on visual inspections and gut feelings. This often leads to inefficient production and loss of profits. The problem is understandable; getting accurate measurements of shrimp using manual and visual methods is a difficult task.

The benefits of smart farm management tools Difficult or not, if shrimp farmers want to improve their processes, increase their yields and maintain efficient and thriving businesses they need to make informed decisions about biomass and feeding. To do this, aquaculture producers are following the lead of


53 agriculture and are turning to technology. Farm management tools, such as the XpertSea Growth Platform, can help because they offer automated, reliable and unbiased production data. Pond management starts with farmers knowing how many shrimp they received in a shipment, which means counting them during the initial stocking period. Knowing this number is important because it will impact the stocking density, feeding regime and the health of the shrimp for the coming months. Using a smart aquaculture device, like the XperCount, allows farmers to automatically and accurately perform their counts. The XperCount is a portable piece of hardware capable of capturing unbiased data about organisms while bypassing the inefficiencies associated with traditional manual methods such as wasted time and inaccurate counts.

Shrimp discovered to be smaller than expected with early monitoring - a second case study During a recent trial of the XperCount on a second shrimp farm in India, the owner found that after four weeks the shrimp in his ponds had not reached the predicted growth. He had expected the shrimp to be around 5 grams in four weeks like his other friends and farmers who bought PLs from the same hatchery. Two things could have caused this: 1. The PLs were stocked much higher

(at 200 per m2) compared to the stocking number at other farms (100 per m2). 2. The feeding did not match the required quantity. The technicians on this farm use blind feeding during the first 30 days. The amount of feed is based on the stocking number and not the growth rate. Ordinarily, information about the growth of the shrimp would not have been available since technicians do not monitor the first month after stocking. It is assumed that monitoring with the XperCount gave them unusual insight into their ponds, allowing them to correct for the slow growth.

gies put reliable data within reach, it is an achievable goal.

References 1. Åsa Strand. 2005. Growth- and Bioenergetic Models and their Application in Aquaculture of Perch (Perca fluviatilis). 2005 2. FAO & UNEP. Feed and Feeding of Fish and Shrimp. 1987.

AFΩ

Reliable data—an achievable goal The challenge for both Suyog Aqua Farms and the farm in India was having access to reliable data. For Suyog’s farm, being able to achieve an optimal FCR proved to be difficult when based on infrequent and erroneous physical observations. Also, according to the FAO, measuring shrimp is more effective than weighing, but measuring shrimp by hand can be tedious and inaccurate. Following in the footsteps of agriculture, shrimp farmers and other aquaculture producers are increasingly turning to big data, analytics and cloud computing to help gather, store and analyze their information. They want to take the guesswork out of shrimp farming and increase their control while reducing waste and increasing profits. As new technolo-

More information

Jason Enlow, XpertSea E: info@xpertsea.com


54

EVENTS 2019 February 12 – 14: International Production & Processing Expo Atlanta, USA http://ippexpo.com/

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Submit your event details to: editor@aquafeed.com April 11 – 12: Aquafeed Food and Feed Drying Technology Course Ås, Norway https://www.fie.com.au/events/dryingnorway

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