Aquafeed Vol 13 Issue 1 2021 by Aquafeed Media, S.L.U.

Vol 13 Issue 1 January 2021

AQUAFEED Advances in processing & formulation An Aquafeed.com publication

OPTIMIZING THE FEED PRODUCTION PROCESS Human biomedical technology for fish Fishmeal alternatives Transparency in feed production Published by: Aquafeed.com LLC. Kailua, Hawaii 96734, USA www.aquafeed.com info@aquafeed.com

AQUACULTURE

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Species-specific solutions for a sustainable and profitable aquaculture At Adisseo, we offer species-specific nutrition and health solutions to aquaculture customers around the world. There is a lot to gain by optimizing your feed additive strategy. Our aqua experts are passionate to help you find out how to increase your productivity and profitability. We look forward to sharing our vision with you!

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AQUAFEED

VOL 13 ISSUE 1 2021

Contents

FISHMEAL-REPLACING FUNCTIONAL INGREDIENT 37 An alternative technology utilizes plant waste to create an amino acid profile, specifically valuable peptides that closely resemble fishmeal.

THE IMPORTANCE OF MOISTURE CONTROL 14

OPTIMIZING THE EXTRUSION PROCESS 17

How to improve the protein ratio and the profitability in feed production processes through digital microwave moisture sensor technology.

Three important factors improve the production process without compromising the nutritional value and feed preference.

NUCLEOTIDES IN FISH NUTRITION 31 Nucleotide supplementation can improve resistance to bacterial and viral infections, the health status of fish in general and reduce ectoparasitic infestation.

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

AQUAFEED

VOL 13 ISSUE 1 2021

Contents 6

Interview

News Review

14 Water underwater?

17 Optimizing the extrusion process to support safety, efficiency and economics

21 The vacuum coating revolution in feed production 24

aking sense of data transparency, M gaining the most from your data

27 Who is accountable? The need for transparency in the feed supply chains

Nucleotides: an important tool for your fish

T ranslating human biomedical technology for fish health and nutrition

Pekilo protein: Past, present and future

A look at the science behind fishmeal-replacing functional ingredient Dietary sodium diformate improves growth performance of the giant freshwater prawn under controlled conditions in the Philippines Fumonisins impact in aquaculture: A real threat?

Columns 29 Gustavo Bozano â&#x20AC;&#x201C; Feed developments in tropical fish and shrimp in South America

47 Albert Tacon â&#x20AC;&#x201C; Nutritional fish & shrimp pathology - I

Calendar of events

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

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

INTERVIEW

with Huang Jie

Huang Jie is the Director General of the Network of Aquaculture Centers in Asia-Pacific (NACA). He was the Principal Investigator of the Maricultural Disease Control and Molecular Pathology Laboratory, Yellow Sea Fisheries Research Institute (YSFRI), Chinese Academy of Fishery Sciences (CAFS), the Chief Scientist of CAFS on aquatic animal disease control, an OIE designated expert for White spot disease (WSD) and Infectious and hematopoietic necrosis (IHHN), and a doctorial tutor for Shanghai Ocean University. Centers and Participating Centers to share their expertise and facilities for mutual benefit. The mission of NACA is to assist members in promoting rural development through sustainable aquaculture and aquatic resources management, which is addressed through five thematic work programs on sustainable farming systems, aquatic animal health and biosecurity, genetics and biodiversity, food safety, quality and certification, emerging global issues, etc. and additional cross-cutting programs including education and training, gender, and information and communications.

AQUAFEED: Would you explain the structure and mission of NACA? HJ: The Network of Aquaculture Centers in Asia-Pacific is an intergovernmental organization established in 1990 after the 14-year implementation of FAO/UNDP projects. Current members are Australia, Bangladesh, Cambodia, China, Hong Kong SAR, India, Indonesia, I.R. Iran, Korea (DPR), Lao PDR, Malaysia, Maldives, Myanmar, Nepal, Pakistan, Philippines, Sri Lanka, Thailand and Vietnam. The core of NACA is a collaborative network of research centers distributed throughout the region. The network is operated by the Secretariat settled in Bangkok and underpinned by Regional Lead

AQUAFEED: What are the main obstacles and solutions to sustainable aquaculture development within the NACA region? HJ: Aquaculture development in Asia-Pacific depends on resources, intensive labor, technologies, and trade and faces the risks of biosecurity, environment, safety, and climate change. The dependence and risks interactively impact the development sustainability. The sector in the region is featured with significant diversities of species, industries, capability, knowledge and culture. Our work’s principal goal is to mobilize the member’s resources, strengthen the region’s connection in aquaculture, and enhance communication and cooperation. To achieve this goal, we plan to develop specific subject-focused networks (sub-networks) to let member institutions in the name of NACA guide the construction of such sub-networks. Member government agencies, industries, and other stakeholders can be involved in the sub-networks. The sub-networks will set up the

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

communication mechanism and promote subjectfocused activities. With the sub-networks, participants can find opportunities for communication, cooperation, training and reaching a consensus on the subject. We believe these efforts will provide regional approaches to reciprocate the dependence, complement the diversities, and minimize the risks and support sustainable aquaculture development. We hope that this idea will be supported and implemented by member governments through the Governing Council. AQUAFEED: Fed aquaculture production has outpaced that of the non-fed subsector in world aquaculture in recent years. Could you give us a regional overview of fed species? HJ: This region is the most diverse aquaculture region in farmed species. A comprehensive enumeration of all species may not be easy. In terms of fish, there are hundreds of fish species such as tilapia, carps, croakers, flatfishes, striped bass, catfishes, groupers, ilsh, Asian seabasses, eels, snakehead, sturgeons, trout, etc. In terms of crustaceans, there are mainly Penaeus vannamei, P. monodon, Macrobrachium rosenbergii, Procambarus clarkii, Chinese mitten crab, swimming crab, and so on. In amphibians, there are mainly bullfrogs and some other farmed frogs. For reptiles, there are soft turtles and turtles. In terms of mollusk, a fed species is abalone. Even sea cucumber of echinoderm can also be a fed species. In general, farmed fish and crustaceans, including tilapia, carps, and P. vannamei, are among the largest dominant fed species in this region. In addition to species, there are differences in life stages and farming methods. Many species during hatching or larval stages and the farming of the above species under high density are feed dependent farming. However, many varieties are not necessarily feed dependent in low-density conditions. AQUAFEED: Many investments are being made in new feed facilities in the NACA region, but what is the actual status of feed availability, and what are the main challenges for feed development in the region? HJ: As the largest aquaculture region, we are also the largest market for aquaculture feed. The region has a large number of feed enterprises on various scales. Feed usually constitutes the largest part of the

production cost of feed-dependent aquaculture. It is also a major factor affecting the growth and health of aquaculture species, as well as the aquaculture environment. Therefore, the quality and cost of feed are the primary concern of aquaculturists. The region has very diverse feed variety and application. We have the most primitive raw feed and self-formulation as feed application, as well as modern full formula feed, including the full formula of broodstock and seed feed, etc. Sources of feed ingredients, formula and processing technologies, additives, pollutants, packaging and storage technology, quality management system (QMS) and other relevant technologies affect the feed productsâ&#x20AC;&#x2122; quality and safety. The marketing system and supply chain of feed enterprises also guide farmersâ&#x20AC;&#x2122; feeding skills and the quality of farming products. Therefore, the market demand for aquatic feed is driven by the huge scale of aquaculture. The quality and technical upgrading of feed products attract wide concern. The feed industry development needs to face challenges in many aspects, such as the rising cost of high-quality fishmeal and fish oil brought by a decline in the wild fish resources, as well as the fluctuation in the quality, supply chain, and price of soybean meal and other major feed ingredients. The feed producers have to face the challenges of achieving the balance of quality and cost of feed products and maintaining the stability of feed quality, safety, and services. Technical innovations and strict QMS will help feed producers achieve their core values to gain advantages in the fierce industrial competition. AQUAFEED: New viral diseases have devastated shrimp production in recent years. What is currently the incidence of viral shrimp diseases, and why is it still having a strong impact on the industry? HJ: In recent years, shrimp aquaculture has faced a variety of disease problems. In addition to the continued impact of white spot syndrome virus (WSSV) on the shrimp farming industry, bacterial acute hepatopancreatic necrosis disease, digestive pathogenic dysbacteriosis, and fungal microsporidia Enterocytozoon hepatopenaei (EHP) have also severely hit the industry in different countries. The newly discovered decapod iridescent virus 1 (DIV1) has begun to pose a threat to the culture of many shrimps

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

such as white leg shrimp P. vannamei and giant freshwater prawn M. rosenbergii. So, it's not just the virus that's causing major losses. NACA established the Quarter Aquatic Animal Diseases Report (QAAD) in this region, and OIE (World Organisation for Animal Health) requires its member countries to report aquatic animal diseases through WAHIS (World Animal Health Information System). However, these reports usually do not request information on the quantitative levels of the impact or occurrence. Some countries may not have comparative surveillance programs or publish their surveillance results. Comprehensive statistics on the incidence of diseases in the whole region may not be available. China has launched a nationwide target surveillance program for important shrimp diseases and annually reported the results in Aquatic Animal Health in China released by the Bureau of Fisheries of Ministry of Agriculture and Rural Affairs and National Fisheries Technology Extension Center. According to the annual report, the positive rates in detected samples in 2018 was 11.7% (WSSV), 10.4% (IHHNV), 12.2% (DIV1), and 22.4% (EHP). Such results would be a reference for the region. There is biosecurity imperfection of shrimp seedlings and farming in this region. First of all, live feed or raw feed is often needed in the breeding of shrimp seedlings, such as silkworms and brine shrimp. Most of these live or raw feed is from farms or areas lacking biosecurity. Secondly, many hatcheries cultivate the broodstock with commercialized subadult shrimp that might be exposed to pathogens in farms lacking biosecurity measures. Third, many native species use the natural broodstock from wild resources or domesticated for a few generations, lacking strict inspection of diseases. Fourth, many farms are densely stocked and lack effective disinfection measures for ponds, water, and wastewater. Fifth, farms carry out polyculture with various crustacean related species, resulting in the spread of the virus among different species and strengthened pathogenicity. These defects make it very easy for known or unknown viruses to enter the farming system. It is worth noting that hundreds of new viruses have been found in crustaceans in recent years. If the above problems are not paid attention to, emerging viral diseases may continue to appear in the future.

AQUAFEED: What are the main strategies and solutions to combat viral pathogens of farmed shrimp? HJ: The principal strategies to combat the viral pathogens of farmed shrimp are the application of the biosecurity concept. Recently, we have joined in the FAOâ&#x20AC;&#x2122;s initiation of PMP/AB (Progressive Management Pathway for improving Aquaculture Biosecurity) and also cooperate with the OIE in the adoption of the new chapter on biosecurity in aquaculture establishment in the OIE Aquatic Animal Health Code to promote the application of the biosecurity concept in the region. PMP/AB will adopt a four-step approach of the strengthened cost-effective management of risks posed by infectious agents in the aquaculture sector with shared public-private responsibilities. For the public sector, a comprehensive set of governance, services, and supports on legislations and regulations on aquaculture with strengthened biosecurity concept, biosecurity related standards, zoning and compartmentation, health certification, national surveillance, reporting, and early warning system, etc. are highly recommended. For the private sector, cost-effective management approaches of the FAO PMP/AB are essential for building a biosecurity system. Accordingly, we cooperate with YSFRI to promote the five-level aquaculture biosecurity grades (ABG) for all types of enterprises. ABG1, namely diagnosis-based treatment, the lowest biosecurity grade, can be applied to extensive small-scale farms. ABG2, surveillance-based prevention, can be applied to most intensive or indoor farms. ABG3, risk analysisbased control, is the integral control under risk analysis guidance with an inadequate traceability system. ABG4, systemic disease freedom, can be applied to breeding centers and broodstock producers. ABG5, official certification, can be applied to disease-free licensed hatcheries or farms. The public and private sectors shall share their responsibility for biosecurity strategies. Smallscale farms would significantly benefit from the sharing responsibility with the national/provincial/ local diagnostic and rapid responding services. The international or regional organizations can provide international or regional recommendations and

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

guidelines, international or regional training, webinar, and consultation on biosecurity and emerging issues. AQUAFEED: The COVID-19 pandemic had a strong impact on the HORECA sector and aquaculture production. What is the impact within the NACA region, and how the industry has responded? HJ: The value chain of aquaculture has been tremendously impacted by the COVID-19 pandemic in the region. As the major aquaculture region globally, the aquaculture in Asia-Pacific deeply relies on the international, regional, and domestic markets, of which a large part is from the demand of the HORECA sector brought by travel and tourism. During the pandemic period, aquaculture products face severe backlogs, and the prices have fallen sharply. Supply chains for aquaculture related commodities, such as seeds, feed, and additives, were also affected. The commodities face difficulty to be accessed by the industry. Aquaculture related services, such as diagnosis, consultation, customer services, etc., have also become hard to access. Labors are hard to find due to movement limitations. Job opportunities have also declined dramatically. Some aquatic products have been banned because of changes in wildlife regulations and the detection of SARS-CoV-2 RNA positives. We analyzed data provided by Fishfirst in China and revealed that shrimp prices in 2020 had decreased by about 25.7% averagely compared to that in the same period of 2019, especially the decreases up to 30-45% in important festivals. Aquatic diseases are also on the rise due to the recovery of farming production following the containment of the COVID-19 outbreak, with an early absence of diagnostic services or inadequate biosecurity measures. If the outbreak crisis threatens food security and causes food exporters to restrict soybean exports, the aquatic feed production will fall short, leading to higher prices. This situation will further lead to higher prices of aquatic products, contraction of markets, and narrowed profit margins. The economies and employment that are primarily dependent on international trade in aquatic products will be significantly affected. The industry also actively responds to the impact of COVID-19. For example, many marketing, trades and services related to aquaculture have been moved online. Online communication and training for

Aquaculture biosecurity grades based on the implementation of biosecurity plan on farm level.

aquaculture technologies have been wildly boosted. Cooperating with our membersâ&#x20AC;&#x2122; resources, NACA has organized or participated in several webinars and training programs for the region and the world. The webinars and training program participants were much more than our earlier physical events, and the countries of the participants are also much wider. Recently, 15 countries in the region have signed the Regional Comprehensive Economic Partnership (RCEP) Agreement. The agreement will ensure that more than 90% of goods to be open to trade. RCEP includes terms of economic and technical cooperation, provides transitional arrangements for the least developed countries and provides favorable conditions for their integration into regional economic integration. I believe that this agreement will bring opportunities to reduce the negative impact caused by the COVID-19 pandemic and promote cooperation in aquaculture. NACA hopes to initiate the concept of aquaesthetics, which means the esthetics in aquaculture and aquatic products. This concept includes the establishment of experiences of eco-friendly aquaculture communities, ornamental aquatic organism industry and marketing, aquatic delicacies, aquatic health cares, etc. The concept will bring new types of aquaculture industries and extend the value chain, attract investments, and attract the general public to enjoy aquaculture. I think the concept will greatly enhance the industry's resilience against COVID-19â&#x20AC;&#x2122;s impacts. AQUAFEED: Asia Pacific countries are among the most vulnerable countries to climate change. Which are the main impacts and efforts to mitigate its effects in the region? HJ: The extreme climates may cause primary disasters, such as typhoons, rainstorms, hyperthermia, drought,

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

etc., or secondary disasters, such as surges, dam collapse, human plagues, social-economic crises, etc. As the infrastructures are inadequate, and farms are usually densely arranged, the primary and secondary disasters may directly or indirectly cause huge damage to aquaculture production. The non-extreme climates, such as the inappropriate temperature, excessive or lack of rainfall, climate fluctuation, etc., may not cause direct damage but deviance and fluctuation in the aquaculture environment. Subsequently, the deviance and fluctuation may cause the decline of growth, disease resistance, the survival of aquaculture species, propagation of pathogens, etc. The efforts to mitigate the impacts should also use the approaches with sharing public-private responsibilities at the farm, local, national, and regional levels. The farm owners shall enhance their infrastructures to strengthen the capability and preparedness facing extreme climates. They shall also establish their biosecurity system and improve the capability of environmental resilience to face the indirect impacts. The aquaculture village/community may enhance their climate services and coordination of areal infrastructures and management shared by farms. Commercial services and product suppliers may develop their services and products to help climate responses, such as forecasting platform, insulation films, escape proofs, etc. The government may establish relevant legislation and regulations on emergency climate responses, establish the climate early warning system, mobilize resources, set up the mechanism for insurance and financial aids for farmers, and support research on climate responding approaches, etc. Regional organizations may establish a network on climate change issues, coordinate the cooperation among countries with sharing water bodies, organize the training for capacity building, etc. These coordinations are included in our strategic plan, and we welcome our members and other organizations in the world to contact us to request their needs and share their resources for the reciprocal development in aquaculture.

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

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

BioMar continues to invest in the shrimp segment The company opened a new extrusion line in the Ecuadorian production facility. This investment is one more step in BioMar’s strategic plan for the shrimp business. The new line will bring the factory close to

200,000 tons of total production capacity. Together with the inauguration, BioMar launched a new high-end diet, EXIA Maxio, targeting the most intensive part of the Ecuadorian shrimp farming sector.

New partnerships to upscale RAS feeds Fredrikstad Seafoods for three more years to develop a sustainable feed recipe for the RAS facility in Fredrikstad. Meanwhile, Green Plains and Hayashikane partnered to deploy feed solutions for RAS systems and beyond. The initial phase of the partnership will focus on Optimal Aquafeed’s deployment of proprietary solutions, developed by Hayashikane, to North and Central American markets.

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Nutreco to expand its learning about RAS and continue Skretting’s development of the feed tailored for RAS systems. BioMar signed a letter of intent with Lighthouse Finance-owned Quality Salmon Sotenäs AB to provide feed solutions for the Atlantic salmon in Quality Salmon’s land-based RAS facility once production commences in 2022 in Sotenäs, Sweden. BioMar Norway also extended its longterm RAS feed agreement with

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Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021 102.13 [2594] 111.12 [2822] 195.72 [4971]

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Salmofood secures license agreement for STIM's smoltification feed solution

Feed Design Lab completes renovation of its pilot plant

Feed Design Lab, a Dutch practical research and education center for

innovation and sustainability in the animal feed industry, completed the renovation of its pilot plant, now fully operational. The center offers a pilot plant to the animal feed industry to perform its own trials or participate in practical training. Partners are also invited to participate in innovative projects, workshops and networking events.

CAT to open aquafeed extrusion facility STIM partnered with Salmofood to supply the Chilean market with its feed-based smoltification solution, SuperSmolt FeedOnly. “We are pleased to have found a professional local partner to produce SuperSmolt FeedOnly for the Chilean salmon industry and look forward to a great collaboration in the years to come,” said Eduardo Hofmann, general manager of STIM Chile.

The Canadian Center for Aquaculture Technologies (CAT) plans to open a new Aqua FeedTech extrusion facility. It will house a complete pilot-scale feed processing line including two feed extruders and a new nutrition analytical lab. The pilot-scale facility will become operational in 2021 and will address specific needs not currently being met, such as producing small batches of research diets needed for

testing novel ingredients, customizing and developing new feed formulas and providing a better understanding of the impact of feed processing on pellet quality.

Innovafeed accelerates its international development InnovaFeed partnered with ADM to deploy its industrial model on the largest American agricultural site (and the largest corn processing site in the world), in Decatur (Illinois). Construction will begin in 2021 and targets a capacity of 60,000 tons of insect protein per year. Meanwhile, InnovaFeed opened the world’s largest insect protein production site in Nesle, North of France with a capacity of 15,000 tons of insect protein.

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

NEW ON THE MARKET Extru-Tech introduces new vertical cooler upgrade option Centered around an advanced feature sanitary cone, a new Vertical Cooler Upgrade Option, recently introduced by Extru-Tech, Inc., promises a new level of food safety and cleanability. While the primary focus of the upgrade was on the ability to provide a safer, more consistent product, customers will also benefit from less cost and downtime for cleaning and improved cooling ability.

Evonik takes NIR technology for raw material and feed analyses mobile

Evonik launched AMINONIR® Portable, a mobile NIR service with a world’s-first feature, amino acid calibrations. AMINONIR® Portable enables the reliable determination of energy, nutrients in feed raw

materials and feed, as well as amino acids in feed raw materials, on-site and independent of a laboratory. The hand-held device connects with the user’s tablet or cell phone. It only requires mobile signal reception and a handful of feed or raw material without further sample preparation to determine their quality at almost any location within minutes. The device and corresponding service packages can be ordered at Evonik's new e-business portal, myAMINO.

BASF, Adifo Software partnership to optimize feed formulation BASF and Adifo Software developed a new digital solution for the animal agriculture value chain by integrating sustainability analytics powered by BASF’s AgBalance® Livestock project into Adifo’s BESTMIX® feed formulation software. By integrating BASF’s sustainability analytics into BESTMIX®, customers along the animal agriculture value chain will be able to strategically manage and optimize feed formulation based on nutrition and cost while taking environmental sustainability aspects into account. Companies hope to commercially launch this innovative solution to the market in early 2021.

FAMSUN introduces new micro-pellet shrimp feed pelleting line The new shrimp feed pelleting line helps producers increase their micro-pellet feed supply during the COVID-19 crisis. The optimized line has proven itself by increasing production capacity from 3-4 t/h to 5-6 t/h when processing pellets as small

as 1.0 mm in some feed mills since May 2020. The improved automation level of the whole pelleting line also helps to reduce operational complexity and allows producers to continue their production with fewer operators in the time of the COVID-19 crisis.

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

Water underwater? Alessandro Mario, Hydronix Fish feed has multiple forms and a wide variety of formulations with different ingredients. In the process, ingredients go through similar steps, including storage, milling, drying, conditioning and pelletizing. Imagine that it would be possible to improve the protein ratio and the profitability in your processes, while saving energy, reducing waste and carbon footprint. Sure, it would be a dream but is it possible? Let us analyze the processes to see where we can find a solution.

Storage The ingredients must be stored and preserved correctly to prevent mycotoxins, spoilage, or heat spots. These often give problems that are directly related to the moisture contents and so monitoring water content while transferring into storage is important to be able to react to problems. Drying Drying is a common practice to store materials safely, and it is a delicate process to reach the perfect moisture target. If you miss the target, the ingredients are prone to mycotoxins and spoilage. On the other hand, over-drying is not just an expensive waste of energy but can cause damage, shrinkage and loss of yield. The material entering the drier has variable water content, and this makes it challenging to regulate the amount of time the material needs to be exposed to the heat or to regulate the temperature. In this process, inline moisture control is used to automate the dryer to save money and improve quality. Conditioning After drying, depending on the material and system requirements, it may be necessary to reintroduce moisture into the product by conditioning. This can be done before milling or pelleting operations. Depending on the final application, the conditioning

Figure1. Moisture control affects the cost and quality of feed products.

can also heat the material to kill germs, cook ingredients and gelatinate starch. In the same way that moisture control enhances the drying phase, it also improves the conditioning process by reacting quickly to changes in the input material moisture.

Grinding Grinding is one of the most energy-consuming transformations in many food processes. Through mechanical action, it reduces the size of feed materials such as grain, seed, fruit and many more to achieve different chemical and microbiological stability. Results vary based on machines and methods used, as well as toughness and moisture of the material processed. The toughness is the ability of a material to resist breakage. Therefore, tougher material will need more mechanical energy to reduce size. The plasticity or ductility of a material determines the amount of energy absorbed before breaking down as well as the final size. More plastic or ductile material will need more energy to break, but it will maintain a more regular final shape. In contrast, less plastic or ductile material will shatter into finer and irregular shards like particles.

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

Table 1. Moisture technologies available in the market.

Type of Technology

Maintenance

NIR/Infrared High

Temp

Salts

Density

Resistive Medium

Capacitive Medium

Microwave Analogue Medium

Hydronix Digital

t Very low t

Nuclear

Very Highs s

Dust

Colour

Note Surface reading

Unreliable

Flowing materials

Only huge quantity

The water content affects the elasticity of the material. Therefore, by controlling the moisture of the material, it is possible to determine the energy consumption of the process, the final size of the powder particles, and the product yield and loss. For these reasons, the initial moisture of many feed materials is the most important element to regulate before the grinding process.

Pelletizing Pelletizing is the process of extruding the formulation into cylindric shapes that are more easily consumed by the fish. The content of the mix is extremely variable between the various applications and formulations. However, even in this process, water content is still an important factor to measure the quality of pellets. Drying pellets The final drying is essential to ensure the pellet shelflife and quality. As explained before, drying is a delicate procedure where it is important to achieve a specific water content, and inline moisture control will help you adjust to the variation in the material. Control and sensors In summary, the moisture affects the costs and the quality of the products. Knowing and subsequently controlling the water content of the material in every step of the process is necessary to improve efficiency, reduce carbon footprint and save money. By controlling the moisture during the process, it is possible to calculate the protein content of the pellet and guarantee superior product quality. To achieve these results, sampling the material is not enough because the samples may not be representative of the full batch, and the speed of the feedback process

is not adequate. It is possible to achieve real-time control in the process with inline sensors. The solution is simple, as using an online sensor makes it possible to know the moisture in the material (Fig. 1). Measuring the water content is an indirect measurement, meaning that it is calculated from another measured property, so to do this accurately, other factors need to be kept as constant as possible: • Material composition • Particles size • Pressure on the sensor • Flow speed • Position of the sensor For this reason, it is key to calibrate the moisture sensors for each material after the final system installation. The calibration must be realized by accurate lab tests, as calibrating any sensor with another different sensor can cause a sum of errors resulting in incorrect calibrations, defeating the objective completely. Independently from the measurement method used in the process, it is critical to completely remove moisture from the sample to reach the dry weight during the lab test, as this is what will be used while calibrating the sensor to define the moisture reported by the sensor.

But are all sensors equal? There are many moisture sensors available on the market, and these different technologies can be

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

Figure 2. Linearity and stability of moisture measurement.

summarized in the five categories in Table 1. An essential but often overlooked difference in the digital microwave technology is the linearity and stability of the measurement. Resistive, capacitive and analogue microwave sensors have a nonlinear measurement making them very difficult to calibrate as they require numerous points to design the curve. Nonlinearity also implies low accuracy at the wet and dry end of the scale (Fig. 2). Sensors with a digital measurement technique have a linear measurement, meaning that the sensor reading and water content are directly and proportionally related. This method allows systems to achieve optimal calibration with a few points. In theory with a linear system, it is possible to achieve calibration with only two points. After these considerations, it is possible to define the ideal requirements of the moisture sensor: • In-line with multiple readings per second, providing quick feedback for the control to adjust every batch. • Robust, made with high-quality materials to withstand tough industry conditions. • Linear measurement, repeatable and stable over time, precise in every condition and simple to calibrate to enable an accurate output. • Store multiple calibrations to be used with different materials. • Ability to measure into the flow of the material.

• Unaffected by dust, color, or variations in salt and mineral content. • Self-contained and easy to integrate into existing systems. • Low maintenance and cost-effective. • Able to monitor and configure remotely for flexible connectivity and analysis. Thanks to the expert research and development team at Hydronix, all the above characteristics can be found in Hydronix microwave sensors which incorporate a unique digital microwave technology made to last in the harshest conditions. Hydronix is driven by the belief that by helping you succeed, we help to build a more sustainable future for our children and the generations to come. For these reasons, Hydronix, with its 39 years of passion and expertise, provides the best digital microwave moisture sensor technology. Hydronix is present in over 80 countries around the globe, providing a network of expert engineers on the field speaking your language.

More information: Alessandro Mario Technical Sales Engineer Hydronix Ltd., UK E: enquiries@hydronix.com

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

Optimizing the extrusion process to support safety, efficiency and economics Arlette Soria, Trouw Nutrition

Manual quality analysis is a mandatory part of aquafeed production.

As the extrusion process exerts intense heat, steam and pressure on ingredients, this phase of aquafeed production is essential to adding functional and organoleptic qualities to aquafeed. Extrusion also presents an opportunity to support aquafeedâ&#x20AC;&#x2122;s safety and shelf-life. Strategic interventions introduced during the extrusion phase can help aquafeed producers achieve safe, high-quality feed, while also contributing to production efficiencies that support producer economics.

However, it is through the extrusion and drying steps the feed undergoes extreme thermal processes that challenge the nutritional balance and feed characteristics. As a result, keeping control of moisture losses, improving production capacity and reducing energy consumption, while safeguarding the nutritional value, can present significant economic savings. This article covers three important factors that can improve the production process without compromising the nutritional value and feed preference.

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

walls, allowing ingredients to destabilize the cell membrane. This mode of action enhances accessibility of the organic acids by decreasing the pathogen cell’s internal pH, inhibiting its growth and eventually killing the mold cell. Along with propionic acid, ActiProp includes a buffering agent to mitigate the acid’s corrosive nature, along with an emulsifier to facilitate intake and spread of acids in the mold cells and phytochemicals. Synergistic effects resulting from the interaction of these ingredients inspired Trouw Nutrition to integrate ActiProp technology into its Fylax Forte HC liquid. Hence, in order to properly select a mold inhibitor, one must be mindful not only to consider the effect of the active ingredients but also the impact the product has on water surface tension. The aim is to find a mold inhibitor with active ingredients capable of prolonging shelf-life effectively by enabling a deeper and stable penetration of the moisture in the kibble.

Figure 1. Propionic acid concentration after extrusion.

Safety and synergy One of the most important quality indicators of feed is the stability of the product for storage. Stability is particularly challenging as the shelf life requirement may reach up to 12 months without jeopardizing important parameters, such as the nutritional value. In order to obtain a high-quality extruded feed, large amounts of water and steam are required throughout the extrusion process. Yet the addition of this moisture can result in the destabilization of the feed in storage given that it provides favorable conditions for the growth of molds, when not managed properly. To effectively mitigate mold growth, a mold inhibitor containing both buffered and non-buffered organic acids should be added at the preconditioning stage. It is important to use a highly concentrated and stable blend that can support high temperatures of the extrusion process and ensure the recovery of propionic acid above 95% (Fig. 1), which help prolonge and ensure the effect of the mold inhibitor. Synergies between organic acids can yield an effect that is more than the sum of individual components. ActiProp is a novel technology developed by Trouw Nutrition that increases the porosity of mold’s cellular

Reducing energy consumption and boosting throughput The different processes occurring during extrusion – such as the large amounts of steam added to increase moisture and the heat required to then dry the kibble – are energy intensive. With the goal of improving efficiency in mind, researchers sought to evaluate if ingredients introduced during extrusion could reduce energy use or accelerate throughput of feed. A validation study was conducted in South America. Researchers added Fylax Forte-HC liquid in two different dosages during the preconditioning stage while maintaining the same production conditions. In other words, the extruder feeder screw speed, the mechanical energy, the steam and water in the preconditioner, the steam in the extruder and the dryer temperature stayed the same. Results, presented in Table 1, reflected a significant increase in the kilograms per hour of feed produced when Fylax Forte-HC liquid was added. The drying process can also require huge allotments of energy. The same validation study found that when the dryer temperature was set to 120°C in all treatments, the feed containing Fylax Forte-HC liquid dried more quickly. These results suggest an opportunity for aquafeed producers to reduce the dryer temperature or increase the throughput speed of feed in the dryer and

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

Table 1. Results of adding Fylax® Forte-HC Liquid to the extrusion process in a commercial plant in South America.

Control Fixed variables

Response variables

0.66 Kg/MT

2 Kg/MT

5.5

9.2

14.3

120

10,000

10,500 11,000

1,100

1,200 1,310

0.4347

0.4321 0.4295

7.9 0.62

thus increase the number of production cycles. Both interventions result in lower energy consumption and/or more efficient production. To further understand the influence on production efficiency, a second study was conducted. In this study, the speed of production was accelerated to 11,000 kilograms per hour and the dryer temperature was set between 105 and 110°C. Again, two treatments were compared with a control treatment. The control treatment contained a dosage of 1 kg/MT of a mold inhibitor previously used by the producer. The two treatments contained different dosages of Fylax Forte-

Fylax® Forte-HC Liquid (Dosage)

7 6.1 0.56

0.43

HC liquid containing the ActiProp (0.66 and 2.00 kg/ MT respectively). Study results, presented in Table 2, demonstrated an improved moisture profile and a shelf life extension of 72% in the treatment with a lower dosage. In the higher dosage treatment, shelf-life was extended 171% compared to the control treatment. The same tendency was found in similar studies conducted in a Latin American production plant with over 1,500 samples – even when the dryer temperature was reduced by 14°C. Studies led researchers to note that time in the dryer could be reduced up to five minutes. And while every production factory is

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

Table 2. Results of adding Fylax® Forte-HC Liquid in finished feed.

Stress test (days)

Moisture (%)

Control

137

7.9

0.62

Fylax® Forte-HC Liquid (Dosage) 0.66 Kg/MT

0.68

2.00 Kg/MT

0.70

for accurate, safe and continuous dosing; and support redundancy by providing back-up equipment. Any manipulation of the production environment must consider its effect on other processes and of course, the final product. Will the finished aquafeed retain its digestibility? Will the characteristics that promote an aquafeed’s sinkage or flotability be maintained – or can they even be improved? Collaboration with stakeholders across the production plant can help assure that adjustments during extrusion result in a product that maintains or even improves its performance.

different, the increased efficiencies could potentially support an additional two-to-four production cycles per day. These studies demonstrate that a strategically selected mold inhibitor can not only inhibit mold growth to support feed safety, but also provide economic benefits via improved energy efficiency and/or greater throughput.

Applying solutions effectively An adequate dosing system is essential to assure the efficacy of a product’s active ingredients. Proper dosing equipment is key to ensuring both effective application and distribution of a mold inhibitor’s ingredients. Due to production variations across equipment and facilities, dosing equipment should be adjustable to the specifics of the production environment. A solution provider should offer dosing equipment that allows

Conclusion As a research-based organization, Trouw Nutrition conducts validation studies at research centers and in commercial production environments around the globe. The studies described above showed that an innovative patented technology science based on carefully blended ingredients and coupled with specific dosing and the right application/distribution equipment can help aquafeed manufacturers achieve consistent quality cycle after production cycle. Strategic interventions can also decrease energy use during the extrusion and drying processes and/or optimize throughput. Ingredient mode of action, careful calibration of temperature and drying times and tailoring processes to the nuances of the production plant and finished aquafeed can support product quality, production efficiencies and producer economics. Photos courtesy of Skretting.

More information: Arlette Soria Nutritionist for South and Central America Trouw Nutrition E: selko.marcom@trouwnutrition.com

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

The vacuum coating revolution in feed production Peter Raeven, Dinnissen Process Technology

Figure 1. A detailed depiction of the vacuum coating process.

A brief history In 1990, vacuum coating techniques were first introduced in the aquafeed production sector by Dinnissen Process Technology. The vacuum coater originated as an evolution of the Dinnissen Pegasus® Paddle Mixer. In mixing experiments carried out under vacuum conditions, high concentrations of liquid were sprayed onto feed pellets. When the vacuum was removed, the liquid was sucked deep into the coated pellets. In this way, Dinnissen succeeded in gradually increasing the fat content of the pellets. Compared to atmospheric systems, up to 80 % extra liquid could be added to pellets coming from the pellet presses. The Pegasus® Vacuum Coater enabled feed producers to apply liquid additives to and into pellets and xtruded products by creating a vacuum environment within the production process. How does it work? Vacuum coating technology allows producers of animal feed to create a vacuum environment within their production processes, enabling them to deal with a wide variety of challenges. The process starts with creating a vacuum inside the vacuum coater where multiple layers of additives can be applied to the product. The Pegasus® Vacuum Coater gently suspends ingredients homogeneously

in the air while the vacuum unit creates a vacuum environment. The spraying functionality makes it possible to spray a precisely predetermined quantity of liquid onto the powders, pellets or granules. When air is then allowed to enter the mixing unit, the liquids are evenly distributed deeply into each particle. With the ingredients being sucked deep into the particles through the vacuum, it also has the effect of protecting them against crumbling. After this step, additional layers of top coatings or aromas are applied to each particle, which results in a high-quality extruded product. Often a thin layer of fat is applied to the granular feed material. This extra protective layer makes the material more elastic so that it breaks or crumbles less quickly. All this is done very fast, since the entire batch process takes just a couple of minutes.

The added value of vacuum coating in feed production After years of developing, optimizing and innovating, the benefits of using vacuum coaters speak for themselves: • More nutritious feed: Preservation of the action of functional additives such as vitamins, minerals, ameliorators, taste enhancers, yeast and enzymes. • Retention of taste and color of feed. • Energy-rich feed via the addition of high

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

Figure 2. Pegasus® vacuum coating system.

percentages of oil or fat results in faster and better growth of animals. • Better feed leads to better digestion, which results in better growth. • Better digestion leads to the reduction of nitrogen, ammonia, phosphate and/or methane emissions (feces). • Healthier animals, and less loss of livestock. • F lexible production processes with sophisticated control programs allow producers to optimize the “recipe” of their product to create an even better feed. • Extremely precise and homogeneous dosing of (expensive) additives, micro-dosing with very high accuracy. • Applicable in different production settings: convenient, easy-to-use and easy-to-clean. • Adding functional additives at the right moment (end-of-line, preconditioning and pressure stage).

Adding functional additives in the final stages of the production process By using vacuum coaters, functional additives can be introduced in the final stages of the production process. Additives, including a wide range of substances such as oils, fats, vitamins, minerals, enzymes, prebiotics

and probiotics, taste enhancers, yeast and other enhancers, can be added to powders, particles and granules. Functional active ingredients can be processed as powders or liquids, and are always added in the vacuum coater after the heating and pressure stages. This functionality of vacuum coaters ensures that heat-sensitive substances remain active after being added to the product. Another advantage of introducing functional additives in the vacuum coater is that the (expensive) additives can be dosed homogeneously and with extreme precision. The vacuum coating process not only preserves the action of functional additives, such as vitamins, minerals, ameliorators, yeast and enzymes, it also helps with retaining the taste and color of the product. An additional advantage of vacuum coating is that pellets aren’t greasy on the outside anymore. This increases the flow out of the silo, prevents pollution and reduces contamination at farms.

The Pegasus® Vacuum Coater vs traditional production methods Vacuum coaters are used primarily for adding essential ingredients such as aromas, oils, vitamins, minerals and enzymes to granules and extruded products. The Pegasus® Vacuum Coater makes it possible to add precisely the right amount of powder or liquid to each granule, after which the additive penetrates deeply into the granule. With the same technology, batches of solid as well as liquid ingredients can be mixed quickly and very homogeneously. The Pegasus® Vacuum Coater is capable of adding much larger quantities of additives to powders, granules and granulates, than traditional production methods.

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

Additionally, it’s possible to quickly add several coatings (one after another) on granules and extruded products and to vary the effect of the vacuum process. For example, switching the vacuum on and off controls the depth to which the liquid additives penetrate into the granule. The sophisticated control program makes it easy for producers to vary the vacuum settings and with it the injection of powders and liquids. As a result, the vacuum coater is suitable for producing a wide range of products on a single production line. Varying with ingredients and switching between recipes is fast and convenient. This also applies to the production of small quantities.

Benefits of the Pegasus® Vacuum Coater for feed producers • Energy-rich, non-sticky and good running granules • Strong pellet, no dust, fewer recalls • Improved efficiency and feed conversion • Lower energy need for production • Possibility of adding components ‘end-of-line’ • Less odor emission, fat/oil is not overheated

• Mixing/blending with vacuum processing capabilities • Highly accurate feeding, weighing and liquid dosing • No contamination and hygienic design for fast and easy cleaning • Suitable for fragile and extruded products • Optimum quality, safety and homogeneity of product, used by many premium brands With a production capacity of up to 30 t/h per unit and batch sizes from 10 up to 3,000 l/batch, Pegasus® Vacuum Coater allows a fast mixing with double shaft paddle mixing technology and a low coefficient of variation. It is fitted with vacuum-tight exclusion valves that have an FDA-approved type seal, developed exclusively by Dinnissen.

More information: Peter Raeven Account Manager Dinnissen Process Technology, The Netherlands E: powtech@dinnissen.com

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100 % natural xanthophylls pigments for greater golden or orange-pink coloring for feeding fish and crustacean

Pellets or crumble

High quality vegetable protein content for improved growth performance

French origin, non GM product, «VLOG geprüft»

DESIALIS - 27/29 rue Chateaubriand 75008 PARIS - + 33 (0)1 42 99 01 01- www.desialis.com - info@desialis.com

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

Making sense of data transparency, gaining the most from your data Paul McKeithan, Bühler Aeroglide

Transparency in food manufacturing is a critical focus for food companies, and feed trends typically follow food. Today, it may not necessarily be a requirement for everyone in the aquafeed industry, but the need for transparency is here to stay. If transparency is a growing business priority, why aren’t more companies focused on it? Perhaps, they don’t know who to ask or what tools to use. Perhaps, they fall into the trap of looking for the latest gizmo, although production of aquafeed is tried and true. Game-changing technology is not that prevalent. Transparency starts with manufacturing operations and data, and has tremendous value when shared and used. While feed manufacturers have a good grasp of their workflow processes, having the ability to make appropriate information available in real-time to regulators, consumers and internal safety personnel is something the industry is still learning about. The idea of “open transparency” is that transparency is in the data, it’s not just about the transparency of the data. The "Open Source Initiative" allows anyone to create modifications to open source

code, port it to new operating systems, share with others and even market it. Although Microsoft first believed it would destroy intellectual property, the company is now the number one provider of open source code. This approach allows natural collaboration and a unique opportunity for the aquafeed industry to partner with open communities. Sharing information can remove points of friction in a supply chain and grow brand equity. In fact, the ability to show transparency may be a processor’s biggest competitive advantage. Forbes called transparency and trust the new currency of brand loyalty. Finding the right partner can help you make sense of data transparency, and help you make the most of your data. By concentrating on transparency, you can become more sustainable in the processes you already know.

Raw material tracing At some point, there will be a need to recall in any operation. Supply chains are increasingly more complex and contain numerous stakeholders, each performing

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

specific roles pertaining to feed production. Raw material traceability means the feed manufacturer has the ability to track a product or ingredient from the point of sale back to its point of origin, with information available about all transactions and movements in between. This can enable the manufacturer to implement a recall in a big cross section of different vendors, even before the market demands it. The roots of traceability stem from food safety concerns and regulations that were pursued to ensure companies had a way to recall contaminated products. Some processors see it as a way to learn what comes into the processing plants and what has left. Some consider it a highly-sophisticated electronic data system, storing product info in barcodes that are easily accessible. Still, others see it as essential information that represents the standards held by a company. All are correct and function to serve the different company and regulatory needs. The type of traceability a company uses will vary and depend on where it is positioned in the supply chain, and the kind of product it handles.

Ingredients and feed certification Global aquaculture production has increased rapidly over recent years, with a larger portion of the world's fish coming from farms, rather than wildcaught sources. As a result, there are widespread concerns about the safety and purity of the resulting products and the effect farming might be having on the environment. The Aquaculture Stewardship Council manages the leading certification and labeling programs for responsible aquaculture, working towards environmental sustainability, while the Global Aquaculture Alliance (GAA) promotes advocacy, education and leadership in responsible aquaculture. In June, the GAA released the latest version of "Best Aquafeed Practices" (BAP), with new guidelines for the selection of ingredients and mill biosecurity. BAP certifies farms, hatcheries, feed mills, processing plants, and pre-processors, based on independent audits that evaluate compliance with the BAP standards developed by GAA. Companies like Skretting already require its aquafeed mills to adhere to certifications

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

and standards. However, certification can only happen when data is made transparent.

Blockchain implementation Blockchain technology is a tool that has the potential to validate the tracking of feed and sustainability data, by linking individual data sets to produce unalterable records and digital ledgers for sharing. Blockchain technology can improve the traceability and safety of feed by creating the history of products and ingredients as they go through the production cycle, especially where environments are complex and fragmented, linking companies and siloed activities. It offers the potential to verify the authenticity of premium products and enhance the traceability of issues that require correction, such as those that lead to product recalls. Many processors perceive blockchain as the key to transparency and how data is shared, but it’s important to realize that this technology is a tool no different than a hammer in a carpenter’s toolbox. Whether or not it is the right tool has yet to be determined. Getting started In most processes in our industry, there’s a correct and understood path to follow, but when it comes to data sharing, there is no standardized process. I’m frequently asked to provide advice, on IoT boxes and gateways, and explain how blockchain is used. The digitalization of data and the idea of transparency is still novel, and there are many available tools. In my talks with the industry, I encourage processors to be curious and open to engaging beyond the equipment, the smart machines, the connectivity platforms and data-sensor boxes. I encourage them to look for a partner who can bring the most value, and in many ways, this is no different from dating. With the right partner, you can determine together the best way to share data. IoT is the enabler, and there is no wrong way to share. Traditionally, a sharing connection might occur once a month by phone. But now, there can be an ongoing connection in the details, with the partner who might ask: How was your day? Can I get you a cup of coffee? Are you picking up the kids, or am I? These connection points are necessary to make the system work. The goal of IoT is to increase these connection points to your solution partners, and it is through these details that sustainability can be achieved.

Finding the right partner Look for the relationship that can offer the most contact points, the most intimacy, and the most shared information, with your ingredient provider, your shipping provider and other connections in the production process. You shouldn’t have to look outside the industry to establish a digital relationship. It can happen with someone you already know. In the initial period, a service will be shaped to discover ways to become more sustainable in the processes you already know. Having the ability to see data in a real-time way, along with your partner, will allow you to see the answers to questions you didn’t know to ask. These won’t be new solutions, but they will offer a more consistent way to execute solutions. Perhaps you operate an aquafeed mill, and the drying process is the final step. When drying parameters change due to temperature or operator error, feed pellets may dry inefficiently. As a result, processing changes must be implemented to get the airflow and bed depth adjusted in order to return to the optimum drying conditions. But what if six months later the same thing happens again. Having real-time transparency with a partner means situations like this can be resolved within minutes, keeping you on task. While your partner stays focused on the drying, watching the real-time data, you can stay focused on other tasks. This is not a new solution, but it is a sustainable way to keep the solution in place using the data you already have. This is not just a computerized dashboard, it is a combination of technology and traditional service with a partner looking at your data, calling and working with you so that you get the most out of that data. When you achieve open transparency in the data, with a partner who will always be there, you will find extraordinary value.

More information: Paul McKeithan, Bühler Head of Digital Services Bühler Aeroglide, USA E: paul.mckeithan@buhlergroup.com

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

Who is accountable? The need for transparency in the feed supply chains Michiel Fransen, Aquaculture Stewardship Council Report of slave labor in Asian forage fisheries, illegal deforestation linked to some Brazilian soy production… (aqua)feed and feed manufacturers are becoming more and more exposed to public sustainability concerns. This often comes with reputational risks – both to the companies involved as to the overall industry. As global attention on sustainability issues increases, it is expected that also public scrutiny on feed-related sustainability issues will rise with it. Supply chain engagement, transparency and therefore accountability, are crucial for its actors in meeting societal expectations to address these concerns. The majority of aquafeed raw material is produced from agricultural crops or animals (aquatic and terrestrial) with the biggest proportion being crop-based (70-75% of the global aquafeed ingredient volume). For decades the aquafeed sustainability discussion has primarily focused on the replacement of marine ingredients with crop-based ingredients. In recent years, this debate has highlighted the need to also focus on social issues in fisheries such as malpractices on fishing vessels, and environmental and social issues linked to poor agricultural practices such as deforestation and community displacement. Given the proportion of feed they already make up, ingredients derived from agriculture are to be considered as additional ingredients, not as alternatives, with respect to sustainability concerns. As for all products that have long and complex supply chains, it is unrealistic to expect the feed manufacturer to have full insight, or control, on matters occurring further down in the supply chain. Ingredients are sourced from global markets and traders, which in turn source from global raw material producers, often linked to other supply chains and subject to multiple processing

steps. This complexity makes product traceability and overall supply chain transparency challenging, and yet, these elements are the “new needs” of supply chains within the wider sustainability discussion. Service-orientated supply chains see these opportunities and are indeed adapting to meet these new market expectations over time. For example, the global coffee and chocolate industry has leveraged a high degree of transparency, traceability and sustainability information transfer within their supply chains. Are these supply chains thus free of problems regarding, for example, social malpractices? Far from it, but while malpractices still occur, greater transparency means the affected actors are able to respond collectively, and thus more effectively, compared with conventional hidden supply chains. As such, supply chain transparency is a means to an end, not an end in itself. Within aquafeed, a good example is continuous progress being made in the marine ingredients supply chain. Persistent efforts by key actors (including major feed manufacturers but also fisheries, processors and traders) have resulted in more and more sustainability concerns being addressed and relevant information becoming available to its users. A pre-competitive attitude has been crucial for this success.

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

Uniquely to the marine ingredients industry is the strong link between marine ingredients manufacturers and aquafeed producers. The same cannot be said with regards “terrestrial” agriculture-derived ingredients. For these commodities, aquafeed is (so-far) a minor part of its end-users. It is thus important to find existing platforms and mechanisms, within these commodities, to collaborate with. Useful examples can be found in the soy and oil palm supply chains for instance. Vital for this transformation towards more transparent supply chains is the involvement of all key actors. It cannot be driven by the feed manufacturers alone. Traders, processors and primary producers must participate and show leadership. After all, if malpractices within the supply chains are found and exposed, it is usually the reputation of the feed manufacturer that faces most public scrutiny on behalf of the entire supply chain. Faced with this dilemma, feed manufacturers rely on either their suppliers to aid in addressing it – or revert to avoid entire supply chains altogether.

Role of third-party certification schemes Across many goods and food industries, third-party certification schemes are used as a means to reduce risks in the supply chain. The Aquaculture Stewardship Council (ASC) is no exception to this. We develop and manage robust standards for responsible aquaculture production, supported by Chain of Custody (CoC) certification that assures consumers that ASC labeled seafood has come from farmers that are minimizing their environmental and social impacts. This allows consumers to reward these farmers, encouraging more producers to improve their practices as well. As part of this program, ASC has developed a separate Feed Standard through a public-facing, multi-stakeholder process. It has taken several years to complete and is expected for release in Spring 2021. The Standard provides a holistic set of sustainability requirements, from feed mill to raw material production, on both environmental and social key impacts. It reaches beyond marine ingredients, soy and oil palm and progresses into additional crop-based raw materials as well. Of special attention is the requirement for feed mills to publicly commit to work towards deforestation-free supply chains, and publicly report on its progress. The Standard not only drives forward efforts to address a number of key sustainability issues for the main raw

material groups but also requires better practices in supply chain management. This is done via extensive Due Diligence and Raw Material Assessment Process for which key environmental and social risk factors have been defined in the Standard. Because transformation and ensuring a gradual transition to higher verifiable levels of “responsibility” takes time for such a complex set of global supply chains, the ASC Feed Standard makes a number of requirements incremental. Based on ASC’s ethos for continuous improvement, a “Marine Ingredients Improvement Ladder” (with different sustainability levels and associated timeframes) and a pathway towards deforestation/conversion-free supply chain are for instance inbuilt in the Standard. A unique core pillar of the overall ASC Programme is transparency, which is central to our efforts to drive up standards and maintain public trust in the seafood industry. For example, all audit reports of all ASCcertified farms are publicly available in full on the ASC website. This aspect is a core element of the ASC Feed Standard as well. Feed manufacturers are required to publicly disclose their supplier Code of Conduct and summaries of Supply Chain Due Diligence outcomes, as well as a number of key parameters, both mill- and ingredient-related. As with the farm audits, feed manufacturers will be subject to independent assessments by accredited and trained auditors with the resulting reports available in-full on the website. Another core Principle of the ASC is collaboration. The challenges facing the seafood industry, and the wider challenges facing the world around sustainability and food supply can only be solved by working together. In this spirit, the ASC Feed Standard offers a unique opportunity for leading supply chain actors to collaborate with feed manufacturers in advancing transparency within the feed supply chain, leading by example and inspiring public confidence in a vitally important industry.

More information: Michiel Fransen Director Standards and Science Department Aquaculture Stewardship Council E: michiel.fransen@asc-aqua.org

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

COLUMN

Regional developments Gustavo Bozano Gustavo Bozano is a consultant at Aqua Lagus. E: aqualagus@gmail.com

Feed developments in tropical fish and shrimp in South America According to FAO data, aquaculture in South America reached a production of 2,716,834 tons in 2018, a growth of almost 80% in the last ten years. Considering the volume produced in that year, 64% corresponds to non-salmonid fish (ten years earlier, this percentage was 57%). Together, Brazil, Ecuador, Chile, Colombia and Peru account for just over 97% of this volume and tilapia, shrimp, mollusks and cachamas correspond to 95% of the species produced by tropical aquaculture on the continent. Two countries stand out for the growth they have been showing in recent years. In 2018, Brazil was already the fifth largest producer of tilapia in the world with 317,080 tons (according to PeixeBr, Brazilian Association of Fish Farming, this number would be just over 400,000 tons) and Ecuador the fifth largest producer of shrimp with 510,000 tons. But does the success of tilapia production in Brazil and shrimp in Ecuador have anything in common? In 1998, Ecuadorian shrimp farmers had reason to celebrate. The volume of production, according to FAO data, reached 144,000 tons, double the volume produced ten years earlier. But between 2000 and 2001, with sanitary problems mainly linked to the WSSV - White Spot Syndrome Virus, Ecuador had a drastic reduction in its production volume. A drop of almost 70% of its annual production, with volumes returning to 45,000 tons, reached the same level as 15 years ago. Without a product to sell and with high production costs, producers suffered huge losses. Several companies closed their doors or were bought by investors who believed that they would overcome their difficulties. In 2016, Brazilian tilapia producers also had a reason to celebrate. According to data from FAO

and PeixeBr, with strong domestic demand, the species' production in ten years jumped to just over 70,000 tons, for more than 300,000 tons. Business was going well and many producers, excited by the good news, continued to invest in increasing their production. But in 2017, the scenario changed. An economic/political crisis hit the country. Fish sales prices, in some regions, were below the production costs of that time. Many producers stopped their production or were bought by companies better structured to deal with the crisis. Both Ecuador and Brazil had their setbacks and, despite different species and problems, found a common denominator for the recovery of their production: the use of nutritional technology to improve zootechnical performance, animal health and the production environment. In Ecuador, in order to overcome the health problems aggravated in 2000/2001, the monitoring and control of water quality, key to the success of any aquaculture activity, has been intensified. Over time, producers began to adopt protocols for the use of probiotics and enzymes directly in the water to reduce organic material deposited at the bottom of the ponds and prevent pathogenic microorganisms. Ensuring a better quality of the productive environment, they acted indirectly in the food and nutrition of the shrimps through the stability in the primary production that also serves as

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

Table 1. The five largest fish producers in South America and their production of the most significant aquaculture groups and species (without salmonids), according to FAO Fishery Statistical Collections for 2018.

Group/species

Brazil

Ecuador

Chile

White shrimp

62,000 510,000 0

Tilapia spp.

317,080 23,050

Mollusks

14,510

Colossoma and Piaractus 157,350 TOTAL

Colombia

Peru

Total

5,397 29,717 607,114

0 77,933 3,075 421,138

10 377,939 0

12,477 404,936

2,867

22,332

182,589

550,940 533,100 377,939 105,662 48,136 1,615,777

food for the animals. In addition, the use of additives in shrimp feed has intensified, improving zootechnical results in production, reducing the amount of organic material released into the water and aiding in animal immunity. Probiotics, organic acids, attractants and immunostimulants started to be part of the daily chat of shrimp farmers. Today, Ecuadorian shrimp lead a sustainable and traceable production model that is an example in the world. Less than five years after the 2001 crisis, its annual shrimp production had already reached levels above that of 1998. In Brazil, during the crisis of 2017, the need to remain competitive in an economically unfavorable scenario was the beginning of a process of intensifying the use of nutrition as a tool to optimize production costs. Brazilian producers were learning that in an environment where the production cycle often lasts more than a year, the permanent adjustment of production strategies to deal with the adversities encountered throughout the cycle was fundamental. Climate, market, political, environmental and social changes require versatility in decision making and oriented nutrition was fundamental for the return to the economic sustainability of the business. Fish production in a country of continental size, with 14 different climatic types, required food producers and manufacturers to structure a nutritional program model that was adaptable to different production systems, in different regions, for different times of the year and with different production goals. It was no longer a matter of designing food only according to the moment of the fish in the cycle (young forms or termination of the animals, or breeding and fry), but a nutritional orientation depending on the particularities of each situation. Concepts related to the benefits of organic acids, probiotics and

prebiotics, flavoring and essential oils in fish feed were disseminated and became part of the daily activity. Today, it is difficult to find a tilapia farm in Brazil that does not use two or three different feeding programs during the production cycle. Using these concepts, Brazilian tilapia farmers were able to optimize their production costs and the volume of tilapia produced in the country has grown again at around 8% per year in the last two years. In 2020, the impact of rising feed costs in aquaculture, due to increases in the prices of the main raw materials used in aquaculture feed, has been a challenge to overcome for producers and feed manufacturers across the continent. Today, aquaculture of tropical species in South America, no matter if it is tilapia in Brazil, shrimp in Ecuador, paiche in Peru or cachama in Colombia, undergo a process of careful evaluation of production costs, which begins with the purchase of the feed. To help producers decide which nutritional program is most effective, some feed manufacturers have been working with spreadsheets to economically simulate the impacts of their different products on production costs and revenues throughout the production cycle. To do so, the understanding of the particularities of each production and the objectives of each producer and the ability to choose the additives that make sense for the feeding strategy, in addition to the knowledge to combine these additives so that there is synergy, without overlapping and nutritional conflicts at a competitive and attractive cost, has been the key for defining the food program to be used and for an assertive orientation. The experiences of Brazil and Ecuador have shown that adjustments and improvements in the efficiency of production processes are crucial for the success of the activity, especially in times of crisis, and that nutrition has a fundamental role in this matter.

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

Nucleotides: an important tool for your fish JoĂŁo Fernando Albers Koch, Biorigin

Figure 1. Recycling system used in the present research.

Nucleotides are considered as precursors of nucleic acids (DNA - deoxyribonucleic acid and RNA - ribonucleic acid) and according to their structure, they can be divided into two main groups: purines and pyrimidines (Fijolek, 2008). In some tissues (especially the liver) nucleotides are created naturally (called endogenous nucleotides) by de novo synthesis. However, exogenous nucleotides (dietary nucleotides) usually are directed to tissues that lack de novo nucleotide synthesis. Examples of this tissue are intestinal cells and also immune cells. Nucleotides are also necessary to recover nucleosides and nitrogenous bases formed during the degradation of DNA and RNA in the salvage pathway (Quan, 1992). Looking at the scientific literature, we can see nucleotide supplementation improving resistance to bacterial and viral infections, health status of fish in general and reducing ectoparasitic infestation (Burrells et al., 2001).

Dietary yeast nucleotides trails Considerable increases in lysozyme activity, serum complements as well as blood neutrophil oxidative

radical production after applying nucleotides in fish diets has been reported for different species. Thus, the purpose of this study was to evaluate the effect of dietary yeast nucleotides (Biotide Extra, 15% of yeast RNA) alone or in combination with HiCell (autolyzed yeast) on growth performance, non-specific immunity and disease resistance in Nile tilapia. Three hundred and sixty healthy tilapia juveniles (20.0 g) were acclimated to experimental conditions for one week prior to the feeding trial. Then, they were randomly distributed as groups of 15 fish into 24 plastic aquaria - 70 L each (8 treatments and 3 replicates) with individual aeration, operated as a closed recirculating system (0.5 L/min) with biological/ mechanical filtration (Fig. 1). Water temperature remained between 22-26oC throughout the trial by conditioning ambient air. During the experimental period, pH was 7.0-7.5, total ammonia-nitrogen was lower than 0.5 mg/L, and dissolved oxygen was not less than 6.0 mg/L. A basal diet was formulated and analyzed to contain, on a dry matter basis, 32% crude protein from meat

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

Table 1. Growth parameters of Nile tilapia fed diets containing different additives (50 days).

Control

Biotide Extra (0,065%) + HiCell (0,27%)

P-value

Final weight (g)

43.2 ± 0.40

46.5 ± 0.60

<0.001

Weight gain (g/fish)

22.83 ± 0.40

26.18 ± 0.60

<0.001

Relative weight gain (%)

112.3 ± 1.9

128.7 ± 2.9

<0.001

Specific growth rate (%/day)

1.51 ± 0.01

1.66 ± 0.02

<0.001

Feed efficiency ratio (g:g)

0.63 ± 0.00

0.70 ± 0.01

<0.001

Final biomass (g)

647.4 ± 6.0

697.7 ± 9.0

0.013

Data are reported as means ± SEM (standard error) of tree replicate tanks. Significant difference between control and treatment was performed using ANOVA and Tukey’s test, P<0.05. Weight gain (g/fish) = (final fish weight) - (initial fish weight); Relative weight gain (%) = [(final weight - initial weight) / (initial weight)] x 100; Specific growth rate (%/day) = (ln(final weight) - ln(initial weight)) x100 / t (days); Feed efficiency (g/g) = (weight gain) / (feed consumed); Final Biomass (g) = Sum of the weights of all animals in the tank.

and bone meal and soybean meal, 10% total lipid from soybean oil. Water was added to the dough and pellets were produced by passing the mash through a 3-mm die plate using a meat grinder. After pelleting, a drying process was performed by forcing ambient air (room temperature) over the pellets for 24 h. Finally, the pellets were broken, sieved to an appropriate size for the fish and stored (4°C) until used. The feeding regime for the trial was 3% of body weight during days 1-40 and 2.5% of body weight during days 41-50. Fish biomass in each replicate tank was determined every 10 days to adjust the feed quantities. Fish received the experimental diets twice a day (9 am and 4 pm). At the end of the feeding trial (50 days), fish from each aquarium were weighed and counted after an overnight fast and the growth parameters were determined (see results in Table 1). Still, blood from seven fish from each treatment was collected with 1.0-mL syringes and was kept on ice up to serum separation (2500 × g for 10 min at 5°C). The immunological parameters analyzed were: production of superoxide anion (O2−) from blood leukocytes (NBT), serum lysozyme activity, serum

myeloperoxidase activity and serum hemolytic activity (complement activity) (see results in Table 2). It is important to highlight that eight treatments were applied in the present research, being composed of associations between concepts. However, the focus of this publication is to present the results of products that contain nucleotides in their composition, such as yeast extract (Biotide Extra – 15% of RNA, source of nucleotides) and HiCell (autolyzed yeast – 6% of RNA). Both products are produced from selected strains of Saccharomyces cerevisiae.

Bacterial challenge After 50 days of the feeding trial, 18 fish from each treatment were randomly selected (from the remained fish) and used in the challenge test. Fish were intraperitoneally challenged with 500 μL of a bacterial solution containing Aeromonas sp. (2.0 OD600). Fish were maintained under the same experimental conditions as reported above (feed management and water quality). Mortality caused by bacterial infection was observed and recorded every 3 hours in each group for 36 hours. The bacterial strain used was previously

Table 2. Immune parameters of Nile tilapia fed diets containing RNA (source of nucleotides).

Control

Biotide Extra (0,13%)

P-value

Respiratory burst

0.397 ± 0.006

0.422 ± 0.005

0.010

Serum lysozyme activity

135.7 ± 19.4

278.5 ± 17.3

<0.001

Data are reported as the mean ± SEM (n = 7 fish/treatment). NBT - production of O2 − (OD 540nm), LYZ – serum lysozyme activity (units/ml). Significant difference between control and treatment performed using ANOVA and Tukey’s (NBT) or Kruskal-Wallis test (LYZ), P<0.05.

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

Figure 2. Mortality of Nile tilapia fed diets containing RNA (source of nucleotides) during 50 days and challenged with Aeromonas sp.

isolated from skin lesions and kidney of naturally diseased tilapia. The total mortality after bacterial challenge can be seen in Figure 2.

Conclusions As nucleotides can be synthesized endogenously and requirements may be met under normal conditions, nucleotides were long considered as non-nutritional. However, both the salvage and de novo pathways are complex multi-stage processes with high energy requirements (Cosgrove, 1998). Moreover, when fish are under stressful conditions (infection, high density or during fast growth and development), de novo synthesis of nucleotides may become limiting and supplementation through the diet can improve both human and animal health and performance, as we could see in this study. Thus, Biotide Extra together with HiCell was able to increase the weight of Nile tilapia fingerlings by more than 3.0 g. In addition, Biotide Extra was able to reduce the mortality of Nile tilapia fingerlings challenged with Aeromonas sp. by 27%, due to the improvement in the defense system demonstrated by the increase in leukocyte respiratory burst and also in lysozyme activity. References available on request.

More information: João Fernando Albers Koch Aquaculture Technical Manager Aquaculture Biorigin, Brazil E: joao.koch@biorigin.net

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Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021 Hydronix-Aquafeed advert-Jan 2021 228.6x58.4mm.indd 1

21/12/2020 13:30

Translating human biomedical technology for fish health and nutrition Marie-Christine Imbert and Timothy A. Bouley, BioFeyn

Better feed for better fish BioFeyn is a new company applying technology developed for human biomedicine to improve fish health and nutrition. Using specialized methods for protecting marine ingredients, this technology amplifies their effect. A system of biocompatible and biodegradable encapsulations is at the core of this science and requires only trace amounts of marine ingredients to be effective. The new method enhances digestion, absorption, bioavailability and delivery to target tissues helping fish grow and remain healthy. This method also has scope to minimize environmental pressures, both in terms of sourcing important ingredients, while reducing pollution associated with nutrient leakage from animal feed and animal mortality-related waste. It is our ultimate intention that this system enables more and healthier fish for farmers, more demand for sustainable ingredients, and more nutrition for a growing fish-consuming population. The encapsulation concept Encapsulation has had a long history in human health and medicine and has been extensively deployed as tool to protect and enable bioactive compounds. Using this technique, nutrients, enzymes, antioxidants, medicines, vaccines and others can be both protected against physical, chemical, and biological stresses and optimized for uptake in the gut and target tissues. The basic premise of protecting and boosting the properties of a “payload” by covering it in another substance works across scale, ranging from nanometers to micrometers and in some instances even at the macroscopic scale.

Figure 1. Negatively charged synthetic polymer shell capsules loaded with curcumin. Table 1. Capsule statistics (from three production batches).

Payload Curcumin

Size (nm) 196.6 ± 6.14

Pdl 0.08 ± 0.04

SEM image.

One recent, prominent example of an encapsulation application at the nanoscale is the COVID-19 vaccine. Both the Pfizer and Moderna formulations each contain mRNA encapsulated in a lipid nanoparticle. These landmark vaccines place a spotlight on this critical area of development for future technologies.

BioFeyn products, insights, and challenges There are many applications of encapsulation technology that can improve aquaculture. BioFeyn solutions, however, focus on protecting the most fragile and limited ingredients. Initially, we have been doing so using nano-scale protections, creating submicron shells and matrices loaded with a compound.

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

Figure 2. Empty positively charged natural polymer shell capsules. Table 1. Capsule statistics (from three production batches).

Payload Empty

Size (nm) 301.5 ± 4.12

Pdl 0.15 ± 0.01

SEM image.

The reasons for operating at this level and with these techniques are manifold. For example, nano-scale encapsulations can: • Extend shelf life • Improve bioavailability (by increasing solubility), protecting from in vivo degradation, enhancing absorption and facilitating intracellular uptake • Control and sustain release • In some instances, target specific tissues/cells Combining these functionalities can lead to orders of magnitude higher potency. This is as pertinent in aquaculture as it is in human medicine. Adapting nano-encapsulation for aquaculture feed faces two primary challenges: (i) encapsulation of

t0: fluorescently-tagged capsule loaded feed pellets versus control.

t60: fluorescently-tagged capsule loaded feed pellets versus control. Figure 3. IVIS Imaging of feed pellets loaded with positively charged natural polymer shell capsules with a fluorescent tag.

relevant ingredients and (ii) integration of these enhanced ingredients into feed itself. Ingredient encapsulation In collaboration with the “Laboratory of Nanotechnology for Precision Medicine” at the Italian Institute of Technology, we have formulated a range of proprietary nanocapsules containing astaxanthin, curcumin, vitamins and other aquafeed relevant ingredients. Intermediary nanocapsules containing fluorescent payloads for imaging were also produced to demonstrate key attributes of our technology. All encapsulation materials have been GRAS (Generally Recognized As Safe) and/or compliant with EU and US feed additive regulation – critical for future market approval of our formulations. Different capsules were successfully optimized from bioderived and synthetic polymers for each payload. We have achieved consistent quality standards with all formulations, as represented by several wellestablished material science parameters: • Target size below 500 nm • Narrow size distribution with a polydispersity index (PdI) of < 0.3 • High encapsulation efficiencies of the compound, even up to 90% in some cases These parameters are very relevant to application in aquafeed. The small target size allows for the nanospecific properties of formulations (in particular bioavailability benefits) to manifest and means that even compounds added to feed at very low concentrations can be uniformly distributed within the feed. The narrow size range has regulatory and stability implications. The encapsulation efficiency is important for cost minimization. Figures 1 and 2 show examples of our nanocapsules under a scanning electron microscope (SEM), with the relevant performance parameters for these formulations summarized in Table 1 and 2 respectively, all indicating uniformity in size and shape. Integration and retention in feed pellets A range of nanocapsules has been successfully integrated and retained in salmonid feed, visualized by fluorescent-tagging of capsules. Biomedical imaging systems (IVIS), typically used for 3D visualizations in preclinical testing, were used to

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

Figure 4. Negligible leakage of fluorescently-tagged nanocapsules from feed.

quantify the integration and retention. Nanocapsulecontaining and control feed pellets were incubated for up to 24 hours in seawater to observe the leakage of capsules. All retention results, even after over 1 hour, were statistically significant. These results clearly demonstrate the suitability of BioFeyn technology for the marine environment and the implications of nutrient retention and therefore animal uptake are self-evident. Figure 3 shows the strong fluorescent signal from the tagged capsules in the feed pellet versus the autofluorescence of the pellet in the control vials at Time 0 (t0, immediately after immersion in seawater) and Time 60 minutes (t60, after 1 hour of incubation in seawater). The results are quantified in Figure 4. These 3D results indicate the even distribution of nano-capsules within the pellet and suggest they do not leak from the pellet within a meaningful time window for aquaculture applications.

Further results Beyond the integration into feed, other analyses of the nanocapsules were undertaken - tests of cytotoxicity, therapeutic potency in vitro, solubility and sustained release profile were all successfully conducted. These results, along with relevant examples of applications in medical and agricultural fields and

collaborations with ingredient producers, allow us to anticipate strong results in upcoming in vivo trials. BioFeyn is aiming for commercial-sized feed batches by the end of the year.

Acknowledgments Paolo Decuzzi, PhD (Italian Institute of Technology) reviewed and contributed to this article. References available on request.

More information: Marie-Christine Imbert CSO BioFeyn Inc., US/France E: m.c.imbert@biofeyn.com

Timothy A. Bouley CEO BioFeyn Inc., US/France E: t.bouley@biofeyn.com

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

PRODUCT FOCUS

A look at the science behind fishmeal-replacing functional ingredient Sara Hash, Menon Renewable Products

As our global population continues to grow, food security and our ability to adequately feed everyone on the planet will be a central concern. While many have discussed the environmental impact of food production and its use of finite resources, something not as openly addressed is the supply chain of feed used in the raising of our protein sources. Of the countless animal feed ingredients, one of the most prevalent is fishmeal due to its nutritional

value. However, fishmeal is not sustainable in the long term given its relatively fixed availability and risk of over-fishing.

A sustainable functional ingredient An alternative technology utilizes plant waste to create an amino acid profile, specifically valuable peptides that closely resemble fishmeal. Menon Renewable Products, Inc. (Menon), an ag-tech startup in San Diego, is one

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

such company working to minimize reliance on fishmeal as a feed ingredient for aquaculture and livestock. Menon has developed a functional ingredient that can eliminate the need for fishmeal while achieving enhanced animal health and improved feed economics. MrFeed速, the feed ingredient developed by Menon, is a disruptive technology that not only replaces fishmeal but has also proved to enhance animal growth and gut health. Through the inclusion of a spectrum of unique prebiotics, nucleotides and peptides, it enhances digestibility and promotes animal health while reducing the need for antibiotics and other disease remediation treatments.

The ingredient not only overcomes antinutritional factors in many animal feed products but also replaces various grains, related proteins, animal byproducts, fishmeal and other components. The product was developed as the result of a project Dr. Suresh Menon was working on almost a decade ago when he was given a contract from the U.S. Department of Defense to hydrolyze agriculture inputs into sugars. His team found they were able to convert these inputs into proteins, lipids and other nutritional components. Following the completion of this project, and seeing the coming challenges in the animal feed industry, Dr. Menon identified a unique opportunity to provide a healthy and sustainable animal feed alternative. This was the motivation behind the foundation of Menon Renewable Products in 2013. With a history in biofuels and cellulosic waste processing, Dr. Menon has expertise in handling organic feedstocks to produce custom-tailored molecules and nutritional components. He was able to develop a patented process that converts hydrocarbon-based sugars from agriculture-based raw materials into a functional animal feed ingredient under the brand MrFeed速. MrFeed速 is a high-quality feed ingredient delivered without artificial preservatives that is rich in essential amino acids. Highly digestible and rich in protein, vitamins and minerals, the product has been developed specifically for use in aquaculture and livestock diets and provides enhanced animal health, superior growth and an improved FCR in order to meet the increasing worldwide demand for an abundant, sustainable, cost-competitive and renewable source of animal feed.

A scalable production process The main differentiating factor between MrFeed速 and other feed ingredients is the structure of the proteins from a molecular level. The size of the peptides can be controlled and make them smaller. As a result,

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

the uptake of these small peptides in the bloodstream is easier, boosting the protein in the blood. This enhances the immune function of a given animal. Through a proprietary and scalable process called CelTherm®, Menon converts hydrocarbon-based sugars from agriculture-based raw materials into an ingredient that utilizes hydrocarbons to create amino acids, fats, nucleotides, peptides and other key components of a functional animal feed ingredient. The Celtherm® process requires optimum levels of hydrocarbon sugars to get the process going. Sugars from untreated feedstocks do not yield much, but after the Menon process of simultaneous saccharification and fermentation, higher levels of saccharified sugars are obtained. The Celtherm® process starts with the breakdown of cellulosic feedstocks into sugars. Over 1,000 unique feedstocks are available in the Menon library to utilize. During the process, the optimal C:N ratio is established, followed by the supercritical processes of oligomerization and optimization of mixing. The Menon team has the ability to optimize sugar levels from a wide variety of feedstocks to customize the ingredient for various animals. Thus MrFeed® Pro50 was developed as part of the MrFeed® line of functional ingredients specifically for shrimp. It is a shrimp feed ingredient with a minimum protein level of 50% and it contains a blend of amino acids, peptides and nucleotides that have been shown to not only promote growth but also enhance digestive health in animals. A similar product is available for finfish. MrFeed® is among the most sustainable options as an alternative for fishmeal. It is currently being produced at scale at prices that are competitive to fishmeal. Menon meets the growing demand for sustainable animal feed utilizing the company’s proprietary process that also reduces stress on the global environment by eliminating fishmeal and other non-sustainable animal feed ingredients. The process is highly scalable and the product lines can be produced from a variety of agricultural inputs, thus alleviating strain on natural resources.

Trial results In addition to the company positioning the ingredient as an alternative to fishmeal, it has also conducted research suggesting that animals fed the product are

healthier and grow faster than those on traditional diets. It has been proven in aquaculture, including shrimp and salmon and other finfish and has demonstrated superior performance when compared to traditional feeds in the areas of survivability, growth, disease remediation, Feed Conversion Ratio (FCR) and overall animal health. In one trial, Menon leveraged the F3 (Future of Fish Feed) Feed Innovation Network (“F3 FIN”). Results from the trial demonstrated that shrimp feed formulations including MrFeed® at an 18% inclusion level, which fully replaced fishmeal, had significantly higher survival rates when exposed to Early Mortality Syndrome (EMS) when compared to the survivability of shrimp fed traditional diets containing fishmeal and fish oil. This proved that MrFeed® can replace fishmeal 100% while achieving significant increases in survivability and growth with overall yields in excess of 25% over traditional diets. In another study conducted with Menon and National Aquaculture Group (Naqua), one of the world’s largest aquaculture operations based in Saudi Arabia, results indicated that shrimp fed diets containing 15% MrFeed® achieved higher survival, greater average body weight and improved feed conversion ratio (FCR) compared to all other test conditions. The study validated the premise that producers can significantly reduce the amount of fishmeal used in shrimp diets. MrFeed® has been tested on more than 400 million shrimp in trials around the world, averaging a 25% increase in overall yield as enhanced immune system health drives faster growth and higher survival. The product has been tested extensively in commercial shrimp ponds worldwide in India, Indonesia, Malaysia, and many others. While additional testing is required, it presents a positive alternative to fishmeal with the potential to grow.

More information: Sara Hash Business Executive Manager Menon Renewable Products, USA E: info@menon.us

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

Dietary sodium diformate improves growth performance of the giant freshwater prawn under controlled conditions in the Philippines Christian Lückstädt, ADDCON

Aquaculture of the giant freshwater prawn Macrobrachium rosenbergii (De Man 1879) started only in the mid 1980s. The latest figures from the FAO reveal that in 2017 more than 260,000 t, worth around $1.5 billion, had been produced – mainly in Asia and the Americas. In the decade leading up to 2002, the growth rate of giant freshwater prawn aquaculture was 31% annually and production is still rapidly expanding, especially in Asia – currently at an estimated annual growth rate of roughly 10%. Macrobrachium is now produced in more than 35 countries over the world. Most of the production of this prawn species is carried out in monoculture in earthen ponds. Commonly used commercial diets include up to 35% crude protein. However, high stocking densities and non-optimal water quality, as well as poor sanitation and non-existent or inadequate quarantine procedures, may impair prawn health and growth performance. Growth may be improved through the application of high-quality feeds, and sustainability of feed ingredient use is one of the main factors for future successful aquaculture operations, among them the use of feed additives that fulfill those demands.

Diformate for the post-AGP era Acidifiers belong to the group of those sustainable aquaculture feed additives. Currently, potassium diformate (KDF), the double salt of formic acid, is

Figure 1. Experimental feed at SEAFDEC Binangonan, Philippines.

widely used in various aquaculture operations, among them in salmon, trout, tilapia, Asian and European sea bass and pangasius. Its value to the shrimp production cycle has also been demonstrated in several field and research trials. The additive decreases gastrointestinal pH and thereby intensifies the release of buffering fluids, containing enzymes, from the hepatopancreas. Formate also diffuses into pathogenic bacteria inside the digestive tract and acidifies their metabolism, leading to bacterial cell death. Furthermore, the available sodium diformate (traded as Formi NDF, ADDCON) has also demonstrated its beneficial impact in several aquaculture species – like trout, European and Asian sea bass, tilapia and carp.

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

Table 1. Growth performance and productivity of M. rosenbergii at 71 days of culture fed with or without 0.5% dietary sodium diformate (NDF).

Control

0.5% NDF

P-level

Final weight (g)

4.1 ± 0.2

4.1 ± 0.1

0.44

Weight gain (g)

3.4 ± 0.2

3.5 ± 0.1

0.42

FCR

2.43 ± 0.19

2.05 ± 0.14

0.02

Survival rate (%)

77 ± 3

87 ± 5

0.01

10.87 ± 1.42

14.70 ± 1.87

0.02

SEAFDEC trial in the Philippines Adding sodium diformate (hereafter referred to as NDF) to the supplemental diet is also expected to improve health and growth performance of the giant freshwater prawn. However, since the available data are scarce – to our knowledge only one trial had been published with dietary acidifier in Macrobrachium so far – a laboratory trial was set up at the South East Asian Fisheries Development Center (SEAFDEC), Binangonan Freshwater Station in Rizal, Philippines. Prawns were kept in tanks, with 15 individuals per tank and four replicate tanks per group in a static renewal system. The size of the tanks was 240 L each. NDF was added to a commercial diet at a dosage of 0.5% (Fig.1), while the diet without NDF served as a negative control. The initial weight of M. rosenbergii was 0.65±0.01 g. Prawns were kept and fed according to normal pond management for 71 days of culture. At the end of the trial, the performance parameters as well as the productivity index have been collected and monitored (Table 1). Only a numerical improvement (+3%) of the weight gain of prawns in the treated group was observed at the end of the trial, whereas the FCR of prawns fed the dietary acidifier improved significantly (P<0.05) by almost 16%. Furthermore, the survival rate – one of the most critical factors in shrimp and prawn grow-out – increased significantly (P=0.01) by 13%. It is, therefore, no surprise that the productivity index, which includes the three most important parameters in every animal production: weight gain, survival rate and feed efficiency (PI = weight gain × survival / (10 × FCR)), was also significantly improved (P<0.05). Here, the improvement was more than 35%, thus leading to a substantially better economic situation, as commercial productivity in shrimp and prawn culture is dominated

by the number of marketable animals and feed costs – and both parameters were greatly enhanced in the group supplied with the dietary acidifier. It should be stated that the impact of the acidifier might have been limited due to the clean laboratory conditions. Effects under commercial pond conditions are often even more pronounced – as this has also been noted for dietary potassium diformate in Vannamei culture.

Conclusion In general, the results show significantly improved performance parameters in giant freshwater prawns fed with sodium diformate. Similar results have already been reported from potassium diformate- (Aquaform, ADDCON) fed white-leg shrimp in Thailand and Ecuador. Therefore, the use of the dietary diformate is supported as a promising alternative in the sustainable aquafeed industry in order to contribute to economic prawn production. Acknowledgments M.L.A. Cuvin-Aralar’s trial work at SEAFDEC Binangonan, Philippines was highly appreciated.

More information: Christian Lückstädt Technical Director FEED ADDCON, Germany E: christian.lueckstaedt@addcon.com

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

Fumonisins impact in aquaculture: A real threat? Rui A. Gonçalves, Lucta

When talking about mycotoxins' impact on aquaculture, the most typical response is “it depends”, which is always a “correct answer”, however, “most of the time” is also an unsatisfactory answer. It is easy to understand that with the high number of produced species (466 species-groups, excluding algae and aquatic plants) in aquaculture, it would be extremely hard to fully understand mycotoxins’ impact in such a highly diverse industry. However, we need to be aware that the extrapolation and/or generalization of the already learned concepts for some species may be incorrect and dangerous.

Fumonisins: What are they? Fumonisins are naturally occurring toxins produced by a number of Fusarium fungi species,

notably F. verticillioides, F. proliferatum and F. nygamai. Several different types of fumonisins are known, but fumonisins B1, B2 and B3 (also named FB1, FB2 and FB3) are the major forms found in feed and food. You will note that most of the experiments performed in animal nutrition tend to use FB1 (purified form) or a combination of all three forms (especially when using natural contaminated ingredients). Fumonisins are characterized by having a long-chain hydrocarbon unit (similar to that of sphingosine and sphinganine), which plays a role in their toxicity. Fumonisins inhibit the sphinganine (sphingosine) N-acyltransferase (ceramide synthase), a key enzyme in the lipid metabolism, resulting in a disruption of this pathway. This enzyme catalyzes the acylation

Figure 1. Fumonisins sensitive levels for freshwater species reported in the scientific literature.

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

and environmental-related (farm management, biosecurity, hygiene, temperature) will highly influence the sensitive levels. Therefore, it is not a surprise that sensitive levels may vary within the same species. Figure 1 shows the minimal sensitivity level for fumonisins reported in the scientific literature for African catfish (Clarias gariepinus), channel catfish (Ictalurus punctatus), African clariid catfish (Heterobranchus longifilis), common carp (Cyprinus carpio), Nile tilapia (Oreochromis niloticus) and rainbow trout (Oncorhynchus mykiss). It is important to Figure 2. Fumonisins sensitive levels for marine species towards fumonisins reported in the scientific literature. note that the EU guidance value for fumonisins (FB1 + of sphinganine in the biosynthesis of sphingolipids FB2) in complementary and complete feedstuffs and also the deacylation of dietary sphingosine and for fish is currently 10,000 µg kg-1 (EC, 2006). As shown in Figure 1, only two species (in this figure) the sphingosine that is released by the degradation have reported sensitive levels below 10 ppm (i.e., of complex sphingolipids (ceramide, sphingomyelin 10,000 µg kg-1) and two reports indicate negative and glycosphingolipid). Therefore, sphingolipids are effects over common carp at the 10 ppm. key components for the membrane and lipoprotein Regarding the African catfish (Clarias gariepinus), structure and will also play an essential role in cell Gbore et al. (2010), reported that the exposure to regulations and communications (second messenger contaminated diets containing 5,000 µg kg-1 or for growth factors). more of FB1, for six weeks, induced a negative What do we know about FUM in aquaculture? physiological response in fingerlings (17.35 ± 1.26 g). In reality, very little is known about the potential Besides negative effects over growth performance, effect of fumonisins on aquatic species. The little FB1 significantly influenced hematocrit, erythrocytes, information available may seem, sometimes, highly hemoglobin, mean corpuscular volume (MCV), variable, even within the same species. However, we mean corpuscular hemoglobin (MCH) and the serum cannot forget that when comparing sensitive levels protein constituents (total protein, albumin and in aquaculture, we may, in many of the cases, be globulin) values. comparing species sensitive levels between a wide In common carp, Pepeljnjak et al. (2003), evaluated range of species with very different ecological and the toxicosis potential of FB1 (500 and 5,000 µg kg-1) in young carps (Cyprinus carpio L.; 120–140 g). biological characteristics. Furthermore, even At both FB1 tested levels, Pepeljnjak et al. (2003) comparing within the same species, it is well observed loss of body weight and higher incidence described that factors as toxin-related (level and of bacterial dermatological lesions erythrodermatitis duration of intake), animal-related (sex, age, cyprini (Aeromonas salmonicida subsp. nova) in the general health, immune status, nutritional standing)

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

group treated with the higher FB1 dose. Several hematological parameters (erythrocyte count, platelet count) and serum chemical concentrations (creatinine, total bilirubin) and activities (aspartate aminotransferase, AST and alanine aminotransferase, ALT) were altered. The author concluded that while not lethal, contamination levels of 500 and 5,000 µg kg-1 FB1 kg-1 body weight could result in adverse physiological effects. In other studies (Petrinec et al., 2004; Kovacic et al., 2009) confirmed that 10,000 µg kg-1 FB1 could affect negatively common carp. The common aspect between all species represented in Figure 1 is their common environment ecosystem, i.e., they are all freshwater species. However, even considering that freshwater species are the most studied group regarding the impact of fumonisins, we are still far from having a full picture of fumonisins negative effects on this group of species. For example, to the present knowledge nothing is known about the potential effect in other important freshwater species, e.g., grass carp (Ctenopharyngodon idellus), the #1 produced species in the world, 10.5% share of all produced species, or silver carp (Hypophthalmichthys molitrix). From the top 5 most-produced species in the world, only common carp and Nile tilapia were studied for the possible impact of fumonisins.

Effect of FUM in marine species Shrimp Fumonisin B1 has not been extensively studied as a shrimp feed contaminant. However, the few studies available suggest that Litopenaeus vannamei is highly sensitive to FB1 (Fig. 2). In 2008, Mexía-Salazar et al. (2008), reported that Litopenaeus vannamei fed FB1 as low as 500 µg kg-1 led to marked histological changes in the hepatopancreas as well as tissue necrosis. This author also found for the first time that both electrophoretic patterns and thermodynamic properties of myosin extracted from shrimp exposed to FB1 were modified, being this an indication that shrimp meat quality was negatively affected. Later in 2013, Garcıa-Morales et al. (2013) re-confirmed Mexía-Salazar et al. (2008) findings for similar contamination levels and further explored the

impact of FB1 in muscle. This study showed that white shrimp fed 600 µg kg-1 FB1 had its growth significantly affected. Interestingly, this author also observed that soluble muscle protein concentration decreased, and changes in myosin thermodynamic properties were observed in shrimp after 30 days of exposure to FB1. Moreover, marked histological changes in the tissue of shrimp fed a diet containing FB1 at 2,000 µg kg-1 were also observed. These findings were further reinforced by showing that FB1 fed animals had a greater decrease in shear forces after 12 days of ice storage, therefore, affecting their shelf life and product quality. To the present knowledge, no information is available about the impact of fumonisins in black tiger shrimp (Penaeus monodon) or any other commercially relevant crustacean species. Fish Regarding marine fish, almost nothing is known about the possible impact of fumonisins in this group of species. Only recently, two studies were published, exploring the impact of this mycotoxin in seabream (Sparus aurata) and turbot (Psetta maxima) (Fig. 2). The gilthead seabream is one of the most important fish species farmed in the Mediterranean. Being a carnivorous fish, traditionally fed high levels of fishmeal and fish oil, its trend to fed with cost-effective plant meals to maintain profitability and sustainability is almost unavoidable. Recently, Gonçalves et al. (2020) reported that fumonisin levels as low as 168 µg kg-1 feed had a significant impact on final body weight, feed conversion ratio and protein efficiency ratio. The authors observed that fumonisins also reduced fat and energy retention and disrupted the respiratory burst in circulating leukocytes. At slighter higher levels of fumonisin (i.e., 333 µg kg-1), these detrimental effects were intensified, clearly demonstrating a dose-response relationship. In other important European carnivorous species, turbot, it was observed that dietary concentrations of 1,000 to 5,000 µg kg-1 of fumonisins in feed, lead to a significant damaging effect on the overall growth performance, nutrient retention, organ morphology and immune status of turbot. Fumonisins tested levels caused a reduction of the brush border villi length in the distal intestine and a reduction of hepatic lipid inclusion (Gonçalves et al., 2020).

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

Conclusions While some freshwater species, namely, channel catfish (Ictalurus punctatus), clariid catfish (Heterobranchus longifilis), Nile tilapia (Oreochromis niloticus) and rainbow trout (Oncorhynchus mykiss) seem to be quite tolerant to fumonisins, others like common carp (Cyprinus carpio) and African catfish (Clarias gariepinus) are apparently more sensitive to FUM. Therefore, not ALL freshwater species may be tolerant to fumonisins. It is important to understand that extrapolation of findings to other species may be dangerous, as we observed within the group of “catfish” sensitive levels between channel catfish and African catfish was impressively different. Very little is known regarding marine aquaculture species, including Pacific white leg shrimp in this group. So far, all data report consistently indicates that marine species might be highly sensitive to fumonisins. Aquaculture is a very diverse industry and will be hard to create guidance values for safe mycotoxin levels in complete feeds that would safeguard all species.

Very restrictive guidance values may be practically challenging with possible important economic negative consequences. However, it is important to further understand the impact of fumonisins in commercially important marine species, especially the species using raw materials with a high risk of fumonisins occurrence. Furthermore, for both fumonisins sensitive and nonsensitive species, it would be very important to study the possibility of carry-over effects. References available on request.

More information: Rui A. Gonçalves Aquaculture Business Developer Lucta, S.A., Spain E: ruialexandre.goncalves@lucta.com

CLEAN FEED. CLEAN WATER. Wenger Extrusion Solutions for RAS Feed Production Wenger innovative extrusion solutions deliver clean, durable, nutritional feeds specifically designed for the most efficient RAS operations. Feeds produced on Wenger systems maintain their integrity better and longer, for clean and clear water. So you feed the fish, not the filter. Learn more about the Wenger RAS advantage. Email us at aquafeed@wenger.com today. PHONE: 785.284.2133 | EMAIL: AQUAFEED@WENGER.COM | WENGER.COM USA

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Nutritional fish & shrimp pathology - I Albert G. Tacon, Ph.D. Dr. Albert Tacon is a Technical Editor at Aquafeed.com and an independent aquaculture feed consultant. E: agjtacon@aquahana.com

Disorders in protein & amino acid nutrition This column is the first of a six-part series dealing with a forthcoming publication by the author dealing with the major reported nutritional disorders in farmed fish and shrimp and represents an update to a previous review published by FAO in 19921. Nutrition is the study of all those biological processes of growth, reproduction and maintenance which depend upon the intake and digestion of food. It follows, therefore, that any negative effects on any of these biological processes can result in nutritional disorders, tissue pathologies and can negatively affect fish and shrimp health. A simplified diagrammatic representation of the possible environmental interactions between nutrition and health in farmed fish and shrimp is shown in Figure 1. Of particular note, is the negative effect of environmental stress and unfavorable culture conditions on feed intake and dietary nutrient requirements, and, in particular, the potential stressful effects of sub-optimal water quality conditions (temperature, oxygen, salinity, natural food availability, pollutants, algal blooms, weather, storm-water runoff, etc.) and sub-optimal rearing conditions (stocking density, water exchange, sediment build-up, human interactions, disease outbreaks, etc.) on the farmed species. Notwithstanding the above, it is generally believed that the majority of these environmental stress factors can be reduced and minimized by creating a more stable culture environment and management system for the farmed species. Notwithstanding the above, for the purposes of this mini-series, the major reported nutritional disorders will be summarized and discussed under six main

categories, namely 1) disorders in protein and amino acid nutrition, 2) disorders in lipid nutrition, 3) disorders in mineral nutrition, 4) disorders in vitamin nutrition, 5) anti-nutritional factors, and 6) feed contaminants and mycotoxins.

Disorders in protein & amino acid nutrition Table 1 shows the reported deficiency signs for individual amino acids when present at sub-optimal levels (i.e. below known dietary requirement levels) within formulated diets for different farmed fish and shrimp species. Of particular significance from the data presented in Table 1 is the important metabolic role played by many amino acids on fish and shrimp health, immunity, gut health and disease resistance, and in particular, the functional amino acids arginine, histidine, isoleucine, methionine, phenylalanine, taurine, threonine, tryptophan and valine. Although specific dietary amino acid deficiencies can be remedied through dietary supplementation with the corresponding limiting free amino acid, it is also apparent from the deficiency signs reported that many amino acids perform other important biological functions over and above to that of just serving as building blocks for proteins and tissue growth (Fig. 2). It follows, therefore, that dietary amino acid requirements for these specific amino acids may be higher when formulating feeds for optimum health and disease resistance, gut health and immunity, stress control and reproductive function. Tacon, A.G.J. (1992). Nutritional fish pathology: morphological signs of nutrient deficiency and toxicity in farmed fish. FAO Technical Paper No. 330, FAO, Rome. 75 pp. 1

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

Table 1. Reported amino acid (AA) deficiency signs and health impacts in farmed fish and shrimp.

Major species - AA

Reported AA deficiency signs & health impacts1

Arginine Channel catfish (Ictalurus puctatus) Golden pompano (Trachinotus ovatus) Jian carp (Cyprinus carpio var. Jian) Largemouth bass (Micropterus salmoides) Nile tilapia (Oreochromis niloticus) Red drum (Sciaenops ocellatus) Red seabream (Pagrus major) Senegalese sole (Solea senegalensis) Striped bass (Morone chrysops Ă&#x2014; saxatilis) Sturgeon (Acipenser schrrenckii x baeri) Turbot (Scophthalmus maximus) Yellow catfish (Pelteobagrus fulvidraco)

Histidine

Atlantic salmon (Salmo salar) Blunt snout bream (Megalobrama amblycephala) Gian tiger prawn (Penaeus monodon) Grass carp (Ctenopharyngodon Idella) Jian carp (Cyprinus carpio var. Jian)

Isoleucine

Jian carp (Cyprinus carpio var. Jian) Olive flounder (Paralichthys olivaceus)

Lysine

Atlantic salmon (Salmo salar) Black seabream (Sparus macrocephalus) Cobia (Rachycentron canadum) Milkfish (Chanos chanos) Rainbow trout (Oncornchus mykiss) Silver perch (Bidyanus bidyanus) Yellow Perch (Perca flavescens) Yellowtail (Seriola quinqueradiata)

Methionine

Arctic charr (Salvelinus alpinus) Atlantic salmon (Salmo salar L.) European seabass (Dicentrarchus labrax) Jian carp (Cyprinus carpio var. Jian) Nile tilapia (Oreochromis niloticus) Yellow catfish (Pelteobagrus fulvidraco)

Phenylalanine

Grass carp (Ctenopharyngodon Idella)

- Reduced disease resistance & immune response - Reduced disease resistance & immune response - Reduced disease resistance & impaired intestinal immune response - Reduced immune response - Reduced disease resistance & immune response - Reduced immune response & intestinal structure - Reduced immune response - Reduced disease resistance & immune response - Reduced immune response & intestinal functionality - Reduced stress control, disease resistance & immune response - Reduced digestive capacity & intestinal morphology - Reduced disease resistance & immune response

- Cataract - Reduced intestinal health & antioxidant capacity - Abnormal histological alterations in the hepatopancreas - Reduced gill structural integrity & antioxidant capacity - Reduced digestive & antioxidant capacity

- Reduced intestinal immune response, tight junctions & microflora - Reduced innate immunity

- Loss of appetite - Reduced hemoglobin level - Reduced hematocrit - Reduced appetite - Caudal fin erosion & increased mortality in fry - Reduced hematocrit & hemoglobin concentration - Reduced sperm quality & motility - Reduced hematocrit

- Cataract - Loss of appetite - Reduced innate immune response - Reduced digestive capacity, microflora & immune response - Reduce immune response - Reduced disease resistance & immune response

- Reduced digestive & immune function & antioxidant capacity

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

Major species - AA

Taurine2

European seabass (Dicentrachus labrax) Hybrid snakehead (Channa maculatus x C. argus) Japanese flounder (Paralichthys olivaceus) Rabbitfish (Siganus canaliculatus) Red sea bream (Pagrus major) Tongue sole (Cynoglossus semilaevis) Totoaba (Totoaba macdonaldi) Turbot (Scophthalmus maximus) Yellow catfish (Pelteobagrus fulvidraco) Yellowtail (Seriola quinqueradiata)

Threonine

Blunt snout bream (Megalobrama amblycephala) Grass carp (Ctenopharyngodon Idella)

Tryptophan

Asian seabass (Lates calcarifer) Ayu (Plecoglossus altivelis) Atlantic cod (Gadus morhua) Brycon (Brycon amazonicus) Chum salmon (Oncorhynchus keta) Common carp (Cyprinus carpio) European seabass (Dicentrarchus labrax) Giant tiger prawn (Penaeus monodon) Grass carp (Ctenopharyngodon idellus) Grouper (Epinephelus coioides) Hybr. catfish (Pelteobagrus vachelli Ă&#x2014; L. longirostris) Jian carp (Cyprinus carpio var. Jian) Mrigal carp (Cirrhinus mrigala) Persian sturgeon (Acipenser persicus) Pikeperch (Sander lucioperca) Rainbow trout (Oncorhynchus mykiss) Totoaba (Totoaba macdonaldi) White shrimp (Litopenaeus vannamei)

Valine

Grass carp (Ctenopharyngodon Idella) Jian carp (Cyprinus carpio var. Jian) Red seabream (Pagrus major)

1 2

Reported AA deficiency signs & health impacts1

- Reduced immune function - Reduced stress resistance - Reduced oxidative stress response - Reduced stress response - Green liver, increased scale loss - Reduced reproductive performance, antioxidant function & larval quality - Green liver, low visceral fat & fat digestibility - Reduced immune function - Reduced immune function, hemoglobin & hematocrit - Green liver

- Reduced immune response, intestinal function & hemoglobin - Reduced intestinal morphology & function

- Increased cannibalism in fry - Reduced maturation - Increased aggressive behavior & reduced stress response - Altered aggressive behavior - Cataract, scoliosis - Reduced stress response - Reduced immune response - Caudal fin erosion - Gill antioxidant system disruption - Increased cannibalism - Reduced appetite, digestive function & antioxidant status - Reduced digestive function - Reduced stress response - Reduced stress & immune response - Increased cannibalism in fry - Reduced disease resistance, stress & immune response - Reduced stress response - Reduced protein digestibility & protease activity

- Impaired gill structural integrity & gill antioxidant status - Reduced digestive function & microbiota - Reduced immune response

Major reported deficiency signs and health impacts in addition to reduced growth and feed efficiency. Non-essential amino acid for most non-carnivorous fish species.

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

Figure 1. Environmental interactions between nutrition and health. Adapted after Waagbo, 1994.

Figure 2. Different roles played by amino acids in the growth, development and health of fish. Adapted after Li et al., 2009.

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

Pekilo protein: Past, present and future Heikki Keskitalo and Simo Ellilä, eniferBio

Figure 1. Pekilo protein.

The past The Pekilo process is an aseptic, continuous fermentation process that produces single-cell protein (SCP). In the process, cell mass with a high crude protein percentage is produced by cultivating mycelium-forming Pekilo fungus in a suitable solution containing carbohydrates and/or organic acids (Forss & Paasinen, 1976). The process was originally developed by the Finnish Pulp and Paper Research Institute (Oy Keskuslaboratorio, KCL), and the process techniques were further developed by Oy Tampella Ab and United Paper Mills Ltd. In all, process development took ten years, from 1963 to 1973 (Ingman, Yrjölä & Forss, 1974). In Finland, Pekilo protein was officially accepted for animal feed purposes in 1971 (Ingman, Yrjölä & Forss,

1974), in 1978 by Czechoslovakia and in 1981 by Sweden (Lehtomäki & Rikkinen, 1982). The first industrial-scale Pekilo process operated from 1975-1982 at the Jämsänkoski Mills of United Paper Mills Ltd, producing Pekilo from spent sulfite liquor, a by-product of papermaking. The production capacity of the Jämsänkoski plant was 10,000 tons of Pekilo product per year, i.e. 10-15 % of the plant’s cellulose production. The second industrial-scale Pekilo plant operated from 1982-1991 at the Mänttä Mills of G.A. Serlachius Ltd. Pekilo contained approximately 50% protein and was used as a substitute for soy meal in feed mixtures for pigs and poultry in Finland. Pekilo protein was extensively tested as an animal feed for pigs and poultry in many countries and was proved to be a safe and valuable source of protein (Järvinen

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

Table 1. Composition of Pekilo protein and soy protein concentrate (g/kg as‐is) and anti-nutritional factor levels in different formats.

Pekilo

SPC

COMPOSITION

Crude protein

650 650

Crude lipid

10 10

Insoluble carbohydrate

220 200 (of which ≥ 50% 1,3-1,6-betaglucan)

Moisture

60 70

Ash

60 70

ANTI-NUTRITIONAL FACTOR

Trypsin Inhibitor mg/g

- <4

Glycinin Antigen, ppm

- <3

b-conglycinin antigen (ppm)

- <3

Lectin (ppm)

- <1

et al. 1980; Kiiskinen & Huida, 1984). In 1979, Pekilo protein was also tested on Atlantic salmon. In the trial, Pekilo protein was used with good results to replace 50% of the fishmeal in the reference diet of Atlantic salmon fry and fingerlings, until the fish were released as 2-year-old smolts (Bergström, 1979). With more than 15 years of safe use in animal feed, no other feed SCP product, with the exception of regular feed yeast can claim the same track record as Pekilo. However, production of Pekilo was abandoned in 1991, due to changes in the Finnish pulp and paper industry that lead to the complete disappearance of the original feedstock – spent sulfite liquor.

The present Many things have changed since the last Pekilo plant was closed in Finland in 1991. Aquaculture has emerged as a major industry to rival land-based animal protein production. At the same time, biorefining processes have evolved globally, and produced several novel types of side streams that could again support the production of Pekilo protein. The founding team behind the new Finnish startup company eniferBio aims to leverage these trends by re-establishing the Pekilo process, now as a major source of protein for the aquaculture industry. The team has validated the process on a wide range of

side streams globally available today. The technology was tested and optimized in the business incubator LaunchPad and hosted by the Technical Research Centre of Finland VTT from 2019-2020. In October 2020, the rights of the original Pekilo strain, Paecilomyces variotii VTT D-75018 / KCL-24, were transferred from VTT to the newly established eniferBio Ltd. Using the newly optimized process, eniferBio is able to generate a product containing 65% protein, which allows it to compete directly with soy protein concentrate (SPC) in feed formulations (Table 1). In addition, the product naturally has a high content of immunostimulatory beta-glucan, which is normally included separately as a premium additive to fish feed (studies on the effect of the Pekilo cell wall beta-glucan on fish gut are currently on-going). The aim of eniferBio is to apply PEKILO® protein in aquaculture feeds for carnivorous species requiring high protein diets, salmon and shrimp in particular. During Q1-Q2 of 2021, eniferBio will perform pilot runs on some of the most relevant industrial side streams and produce hundreds of kilos of PEKILO® protein for extensive fish feeding trials. The company was awarded the first prize in the 2020 edition of the Nutreco Feed Tech Challenge, which includes a scientific validation by Nutreco’s aquaculture division Skretting.

Aquafeed: Advances in Processing & Formulation Vol 13 Issue 1 2021

The future The aquaculture industry is actively searching for alternative feed proteins that could help in maintaining sustainable growth in the sector. The industry has gone through two alternatives in tackling the fish feed protein challenge: originally using wild fish in the form of fishmeal, which then was gradually replaced in an ever greater part by soy protein concentrate. These solutions alone are not viable in the long run. Fishmeal production cannot grow because the global wild catch is already overexploited. Soy, on the other hand, is one of the main drivers of the ongoing destruction in the Brazilian Amazon. The Amazon is vital to the Earth's carbon cycle, storing more carbon than any other terrestrial ecosystem. The Amazon is projected reach its tipping point at which it would flip to a savannah-like ecosystem and billions of tons of carbon dioxide could be emitted into the atmosphere (Amigo, 2020). The Paris accord is impossible to implement if we do not stop deforestation in the Amazon. Aquaculture needs a new solution to meet its protein demand. Recently, two new “alternative proteins” have been taken to industrial scale, insects and methanotrophic bacteria. While potentially sustainable, insect protein cannot compete on price with soy

(Arru et al., 2019). On the other hand, using bacteria to convert fossil fuel (natural gas) into fish feed is not an environmentally sound solution (Le Page, 2016). Any solution that could significantly impact the aquafeed protein landscape would need to be sustainable, scalable, and cost-competitive with soy. The Pekilo process’s industrial history and its ability to derive protein from existing biorefining processes without additional inputs can precisely enable this. One Pekilo plant with a footprint of 900 m2 can produce the equivalent amount of protein as 65 square kilometers of soybean fields. Europe’s bioethanol plants alone produce some 8,500 million liters of ethanol annually (Flach et al., 2019). On average, distilling a liter of ethanol leaves behind 10 liters of side streams in the form of vinasse/stillage, potentially supporting the production of 850 kT of 65% PEKILO® protein in Europe. For comparison, Norway´s salmon farms currently utilize approximately 300 kT of soy protein concentrate (SPC) annually (Aas, Ytrestøyl & Åsgård, 2019). Producing protein for Europe’s aquaculture industry locally, sustainably and economically is possible. References available on request.

More information: Heikki Keskitalo CTO eniferBio, Finland E: heikki.keskitalo@eniferbio.fi

Simo Ellilä CEO eniferBio, Finland E: simo.ellila@eniferbio.fi

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