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Animal feed certification instrument and procedure to guarantee the quality of meat and dairy products through automatic, simple and rapid DNA barcode method based on tubulin-based polymorphism (TBP)

Final Report Summary - FEED-CODE (Animal feed certification instrument and procedure to guarantee the quality of meat and dairy products through automatic, simple and rapid DNA barcode method based on tubulin-based polymorphism (TBP))

Executive Summary:
The FEED-CODE project has achieved its most ambitious expectation to represent a new method for certifying the qualitative and quantitative content of animal feed samples. Consistency and reliability of the FEED-CODE qualitative and quantitative data have been eventually verified in a couple of inter-laboratory comparisons. The FEED-CODE method was shown to be superior to the currently ongoing method for feed authentication that is based on the use of the optical microscope. On the contrary of the later, no false positive, neither false negative results were ever produced by the FEED-CODE method and the estimated weight/weight percentage of single ingredients present in the range of 1-10% was the most accurate. Incidentally, these assays have clearly shown all the weakness inherent to a method that, like the OM's, relies on the individual experience and eye.
The way it has eventually worked out makes FEED-CODE a modular system capable, in response to different needs and requests, of providing either qualitative or quantitative data, or both. In fact, the FEED-CODE system is structured in 4 different operative modules consisting of sample preparation, qualitative analysis, quantitative analysis and data analysis, this later supported by specifically tailored software programs.
In particular, three software applications for the FEED-CODE system were designed and developed: a management software will assist the operator to integrate all the automated and manual steps through the whole procedure (FEED-CODE manager), whilst two analytical software (one for the qualitative data and one for the quantitative analysis), will provide the final output of data analysis, also in the form of a QR code. These SW components were tested individually and integrated in the prototype pipeline.
The qualitative analysis that is the recognition of any botanical ingredient, declared or not declared, is based upon a unique feature that makes FEED-CODE capable of profiling all the DNA molecules of different botanical origin present in any given sample. This same feature turned out to be quite handy and precious for the evaluation of the level of purity of the raw materials used to produce compound feed. The quantitative analysis, based upon the development and use of species-specific probes, all derived from the same innovative strategy of production, has been worked out at metrological standard with the use of reference mixes, unavailable in the field, and up-to the purpose species-specific DNA calibrators, so that the presence of any specific ingredient in a feed sample can be referred to an absolute number, that is the target copy number (TCN). Making use of the reference mix, the w/w percentage of a single ingredient can then be confidentially established, as successfully shown in both ILCs. Indeed, a proper validation of the analytical modules developed was concluded with an inter-laboratory comparison (ILC), organized by Eurofins as a third part, and involving laboratories providing microscopic feed analysis. Results of the blind analysis of seven compound feed-like mixtures gave final credit to the project results. On average, DNA-based methods provided better performance than microscope analysis, and in particular, TBP-based methods were the best performant both at the qualitative and quantitative level.
At present the FEED-CODE combinatorial system, qualitative and quantitative, of feed analysis has been worked out for the qualitative detection of 12 plant species and the simultaneous quantitative detection, in the 1-10% range, of 7 of them that are either those supplied in low amount for nutritional purposes (flax, sunflower or sugar beet) or prohibited by some cheese-specific disciplinary of production (cotton, rapeseed, peanut and rice). This prototype has been brought to TRL9, ready for the market. To this scope, a valuable exploitation strategy and commercialization plan have been assessed and defined with the aim to facilitate its market uptake and therefore the adoption of FEED-CODE system in the reference application domain. Nonetheless, the generation of a database containing the specific profile of 31 different plant species, selected from the EU catalogue, together with the availability of an additional database containing the necessary sequence information of all the introns of the same 31 plant species, makes it feasible to upgrade the FEED-CODE prototype at will.

Refer to the Section 4.1.1 of pdf file for a more detailed description of the Executive Summary and related figures.
Project Context and Objectives:
Project Context
The issue addressed by the FEED-CODE project concerns the qualitative and quantitative determination of the different botanical species, declared or not declared, present in compound feeds, produced and sold in the reference market. The industrial, regulatory and scientific scenarios are briefly described as it follows.
The EU integrated approach, aiming to assure a high level of food safety within the European Union, needs effective control systems to assure compliance with EU standards in the food safety and quality, all along the food chain. Unfortunately, this is not always straightforward, in particular when several actors are involved in the chain, in some cases not even belonging to the European Union. Indeed, although thousands of different compound feeds are produced and commercialized in the EU, most of them relies on raw materials coming from extra-EU countries that must be inspected at the frontier. Animal feed market is huge and valuable: this favours fraudulent admixtures or omissions in feed formulation. To this scope, the level of botanical purity of raw materials is established by EU regulations (Council Directive 96/25/EC). EU regulation defines new “rules on the placing on the market and use of feed for both food-producing and non-food producing animals within the Community, including requirements for labelling, packaging and presentation” and states that all feed material from which animal compound feed are composed must be reported in the label in descending order by weight (Regulation EC 767/2009, Art.17). Percentage by weight of the feed materials contained in compound feed for food-producing animals must be made available to the purchaser, on request.
At the European level, no reference method has been established for the determination of feed botanical composition. The only available method is based on optical microscopy (OM), utilized by few laboratories at the EU level. This is the official method in Italy (G.U n° 123, 28/5/1994, Suppl.11) but it is not an accredited or standardized method. DNA-based analytical methods for food analysis are in use and some of them has recently become the official or recommended methods in the EU: for GMO detection (European Network of GMO laboratories), horse-meat detection in bovine meat (EURL-AP recommended protocol, 2013) and ruminant DNA detection in feed (EU 51/2013).
In this context, the final goal of the FEED-CODE project was to provide a code that certifies and documents the composition of compound feeds in an easy, reliable, fast, affordable and comprehensible way.
In accordance with the EU legislation every kind of feed, as well as food, should be correctly labelled and its composition duly recorded ensuring quality and safety of animal productions and a fair trade. Feed producers could take advantages of FEED-CODE project by certifying their productions while end-users, that may be either producers or consumers of milk/cheese and meat, could find in FEED-CODE a reliable guarantee for their purchases.
A more accurate method of detection of the real composition of animal feed than that relying on OM is largely overdue since this technique suffers of major limitations as admitted by the same stakeholders and ascertained in the course of the FEED-CODE project. In fact, REA has attributed to the FEED-CODE approach: “the potential to clear-off all the uncertainties concerning feed production and consumption by giving the European milk and meat production sector, as well as other sectors, a tremendous boost and an international added value”. Moreover, as stated in the assessment report: “it will better support PDO product qualifications and assurance, as it can provide to any producer or farmer that wants to certify or trace the feed a cost effective means for doing so...”.
On the scientific and technical side, the FEED-CODE project is based on an innovative, specific yet versatile molecular biology approach that allows the concomitant detection of different plant species in the same mixture, the identification of undeclared, contaminating species in both raw materials and compound feeds and the preparation of unique species-specific molecular probes to be used in quantitative assays.
This approach is based on the instrumental use of both length and sequence DNA polymorphisms that are commonly present in the introns of the genes belonging to the plant beta-tubulin families. That is why the method is named TBP for Tubulin-Based Polymorphism. TBP, that is the method on which the whole FEED-CODE approach relies on, is protected by the European patent n. 1144691 and the European pending patent n 11711998 both made available to the FEED-CODE consortium by the Italian National Research Council (CNR). At present no other molecular biology approach can provide, as TBP does, the DNA profiling of all the botanical species, declared or not, actually present in any given feed sample. A further advantage attributable to the TBP-based DNA profiling is the possibility of assessing the level of purity of the raw materials used for making compound feed. Referring to the quantification of the single botanical species present in feed samples, TBP offers the unique possibility of isolating species-specific molecular probes in a rather straightforward and systematic way allowing, at the same time, the convenient preparation of single-species DNA calibrator plasmids.
The applicability in the project has been assessed, with the contribution of SMEs and Associations, by defining the scenario of application, the user requirements and the system specifications. In accordance, FEED-CODE is expected to be useful for:
➢ controlling raw materials botanical purity;
➢ controlling compound feed authenticity and conformity to the label;
➢ respecting PDO production guidelines,
➢ controlling feed production lines,
➢ providing product certification to different kind of feeds supplied to different livestock.

These expectations were also backed up by REA that, in the assessment report, stated that: “the FEED-CODE project has a high chance of generating an important impact at European level, affecting not only the consortium participants but also the different application sectors of interests. It might lead to a new (de-facto) standard for the labeling and certification of feed in support of implementation and application of the regulation EC 767/2009. Furthermore, it was predicted that: “the impact to the level of consumers will be relevant since the perception that the chain of food production is entirely controlled starting from the diet of the animal is a very positive and strong”.
The query of a well assorted group of farmers, compound feed manufactures, milk and meat producers, emphasized their interest on FEED-CODE for certification of feed and traceability, identification of frauds, exclusion of contaminations from undesired species and in compliance with specific production protocols (i.e. PDO products).
Project Objectives
The main objective of the FEED-CODE project was to develop a reliable, simple and affordable technology for the fingerprinting of each individual plant species used for common feed production (species included in the EC Regulation n. 575/2011) quantifying the presence of individual components even in case of very small amounts. This purpose mainly comes from the expressed needs gathered from the stakeholders’ target groups, requiring for:
1. Feed certification for quality assurance of high quality products derived from food producing animals (milk, meat and their derived). Specifically in the case of PDO products, the target is the verification of the application of the specified production protocols.
2. Certification of compound feed mixtures composition and claims, carried out by feed manufacturers.

This objective had to be achieved by combining the two major branches of the whole analytical process capable of providing qualitative and quantitative data from the same feed sample, respectively (Figure 2).
Originally, the whole pipeline was intended to be automatized and assembled in an exclusive piece of hardware, meant as an unique FEED-CODE instrument. However, by proceeding with the experimental work, it became clear that such an achievement was far from the possibility and available resources of the FEEDCODE consortium and, thanks also to REA suggestions made at the interim review meeting, the objectives and aims of the project were more properly refocused towards the generation of a FEED-CODE analysis software to be used with either the analytical CE-TBP platform or the FEED-CODE quantitative array plate.
In accordance, a FEED-CODE manager software capable of recording and organizing each step of the procedure had to be developed, as well as a software suitable for the analysis of the data produced by both the qualitative FEED-CODE platform and the quantitative FFED-CODE probe branches. Technical innovations allowing the processing and the analyses of multiple feed samples had to be also developed.
An additional objective set up at the interim review meeting was the generation of an analytical working prototype targeted to the qualitative and quantitative determination of a well selected set of plant species to be exploited at dissemination level and as a proof of principle, such as: “an assay plate limited to the detection and quantification of few species, but upgradeable based on the RTD's results generated to cover a complete palette of species relevant for an agricultural/farming solution”. The data obtained through the assay plate had to be consistent in terms of presence /absence of specific components with those resulting from the FEED-CODE CE-TBP qualitative analysis.
Given all this, the main original objectives laid down in the DoW needed to be partially revisited for some points of the following original lines:
1. An application scenario and a list of user requirements have to be obtained from stakeholders (original D1.1) providing the actual start of the experimental work.
2. A priority list of the plant species of interest and system specifications ,such as limit of detection, resolution level, repeatability of the data, time of measure, instruments and SW specifications had to be defined (original D1.2 and MS1).
3. A specific CE-TBP profile for each of the 31 selected plant species had to be obtained. This was to establish a FEED-CODE data base made-up by single species standards of reference (D2.1 and MS2).
4. Suitable, consistent and convenient protocols and assays for DNA extraction, DNA purification and for testing the presence and the interference of inhibitory substances had to be worked out (original D2.2).
5. Original CE-TBP protocols had to be implemented with respect to peaks resolution, to the limit of detection, to the identification of diagnostic peaks, to the range of analyzed DNA amounts while using the new DNA analyzer bought with the project (original D2.3).
6. Species specific molecular probes and selective PCR reactions had to be set up (original D2.4 and D2.5). As already anticipated and in accordance with the recommendations of the interim review meeting, this objective was restricted, for the purposes of FEED-CODE prototype, to the 7 plant specific probes of the requested multi assay plate.
7. The limit of single species quantitative detection with CE-TBP had to be determined (original D3.1).
8. Validation at end-point PCR level of the CE-TBP qualitative data by confirmation with either AOX- or TBP-derived molecular probes had to be achieved (original D3.2 and MS4).
9. Detection of single plant species by q-PCR assays based on the use of TBP- and AOX-generated molecular probes had to be shown (original D3.3 and MS3).
10. Single plant species detection carried out with alternative procedures based on microchips or beads has to be demonstrated (original D3.4).
11. A prototype version of the FEED-CODE platform and probes had to be set up and validated (new D3.5).
12. A layout and measure flow sheet of the FEED-CODE operating procedures had to be worked out (original D4.1).
13. Component selection and block diagram of the whole FEED-CODE procedure had to be work out (original D5.1). Moreover, in accordance to the requests received during the interim review “to make the method, tool independent in order to support that FEED-CODE becoming a de-facto standard”, an additional documentation was provided including requirements and recommendations for FEED-CODE automation (new D5.3).
14. Software programmes for data analysis and for automation had to de defined for the FEED-CODE prototype (original D5.2 and MS5).
15. Validation of FEED-CODE system through the implementation of benchmark tests with reference methods and related user acceptance had to be done and verified (original D6.1 and MS6).
16. Feasibility study was carried with the aim to assess the economic sustainability of FEED-CODE technology solution, and related services/products offer (original D6.2).
17. Study of an application model and an analysis of regulations compliance was performed with the aim to define a new standard proposition for animal feed labelling procedure (D7.1 D7.2 and MS7).
18. Creation of proper and efficient communication and training tools (D8.1 D8.2 and D8.4) together with the definition of a proper dissemination plan have facilitate the promotion activity of the main project findings toward a well selected targeted audience of potential stakeholders (D8.3 and D8.5).
19. A proper market analysis and technology assessment was carried with the aim to verify the innovative and competitive level of the FEED-CODE solution proposed, whilst the definition of valuable exploitation strategy and related business model will facilitate the future market uptakes of results and their adoption in the reference value chain (D8.5 and new D8.6).

Refer to the Section 4.1.2 of pdf file for a more detailed description of project context and objectives, and related figures.
Project Results:
Specific plant species probes
All the objectives and the related deliverables and milestones enlisted in the partially revised version reported above, consistent with the remarks, suggestions and requests made in the first assessment report, have been brought to completion. As a service, the FEED-CODE technology is now available for producers and consumers (TRL9) whereas products, like assays kits, are ready for a first of a kind commercial system (TRL8).
This applies to the FEED-CODE prototype that has been developed for the recognition of 12 selected plant species and for the simultaneous quantification of 7 of them, representing, according to the prototype design, either species important for animal nutrition, added in low abundance, or species that are forbidden by some disciplinary of production enforced for cheese makers. As requested, the FEED-CODE combined system of qualitative and quantitative analysis has been brought to a high level of reliability for detection and measurements and is now upgradeable by anyone of the remaining plants species of the project.
More details can be found in the following list that, just for space constraints and sake of clarity is restricted to the major achievements of the projects. The list is proposed in accordance with the following operating procedures, resulting from the application of the workflow depicted in Figure 2.
In accordance to points 1-3 of the previous list of objectives (corresponding to D1.1 D1.2 and D2.1) an exclusive database of the CE-TBP profile of 31 different plants species and some varieties therein has been realized. Plant species were selected from the European catalogue of raw materials used for the preparation of compound feed on the basis of AGs and SMEs requirements. Such a database can be implemented anytime with a practically unlimited number of plant species. This database allows the identification of species-specific diagnostic peaks, very useful for their immediate recognition in the output of the analysis.

Refer to the Section of pdf file for a more detailed description of result n°1: specific plant species probes, and related figures.

In accordance to point 12 of the list of objectives (original D4.1) a layout and measure flow sheet of the FEED-CODE operating procedures has been worked out, including all the operations (cross contamination from powder dispersion, DNA extraction protocols, control of inhibitors, PCR reaction conditions) required to have the system working. As also already anticipated, and clearly stated at the Interim Review meeting, the final layout did not coincide with the construction of a single FEED-CODE instruments but with the setting of a modular FEED-CODE method that is made up by 4 different operative phases.
In this context, the FEED-CODE processes were analysis per each step identified, and each of them is composed of a list of actions that constitute the laboratory protocol defined for FEED-CODE process (refer to Figure 23).
To this respect, and with the purpose to meet the recommendation received by the REA, the Consortium was required to perform additional work to make the adopted method, tool independent: “to support FEED-CODE becoming a de-facto standard”. With this aim, a proper new deliverable (D5.3) was developed in order to provide requirements for FEED-CODE Platform and Probes processes automation. Indeed, going through this procedure manually, means to spend a lot of time (hours/days) to achieve results for a small number of samples that could be affected by human errors whose probability to happen is higher due to high number of manual passages. Both processes are constituted of a multi-step of physical-chemical transformations of variable length in terms of time: starting from the sample to the final data output which constitute the result of the FEED-CODE analysis, the process development itself through an extensive and laborious list of actions which following a specific laboratory procedure.
In this view, three automation scenarios were identified and analysis according to their complexity and cost (each of them can be considered as the upgrade of the previous one):
1st level of automation: it is the level of automation that has been achieved during this last timeframe of FEED-CODE project, mainly making use of standard laboratory equipment. Some high-tech machines have been associated to cover those phases considered high skilled demand and very high time consuming if implemented manually by an operator, whom role is to carry out the sample preparation between each step of the chain of process.
2nd level of automation: the difference between the 1st and the 2nd level of automation is that the sample preparation between phases is also managed by means of high-tech automatic equipment, mainly liquid handling robots. The role of the operator is reduced to load/unload a given machine at a given time in order to proceed with the process.
3rd level of automation: the last level of automation removes completely the role of the operator. The machines selected to carry out the FEED-CODE process become integral part of a full-automated multisystem, which takes as input the animal feed samples to be analysed and to provide as output the result of the analyses.

Refer to the Section of pdf file for a more detailed description of result n°2: FEED-CODE instrument, and related figures.

FEED-CODE certification procedure and service
FEED-CODE certification procedure and service is focused in WP7 activities, including an end-user survey on obstacles (T7.1) and the regulation compliance and new standard proposition (T7.2).
The main objectives of WP7 have included:
• to perform an end user needs survey coordinated by the SME-AGs in order to identify current obstacles decisive for today’s lack of regulatory compliance. This work was carried out in cooperation with the Associations through contacts with their members. In particular, the legal, financial, administrative framework were assessed in order to identify the main barriers to the diffusion and adoption of such technology. Also, the social aspects related to the low tendency of innovation in the quite conservative realities of agro-food sectors have been considered. TI provide support on the regulatory/certification aspects and into the roll out of the survey.
• following the results of the testing/validation in WP6, the compliance with EU regulation with respect to animal feed labelling and the relative certification procedures was verified. Moreover, thank to the expected enhancements of the detection capacity, a new standard for the labelling and the certification will be proposed, as an input to the “EU Code of good labelling practice for compound feed for food producing animals”; including suggestions and indications about potential improvements to the regulation EC 767/2009, achievable through the adoption and use of FEED-CODE instrument and procedure.
In accordance to the point 17 of the list of objectives (D7.1 D7.2 and MS7), Teknologisk Institutt AS (TI) has, with assistance and input from Labor and IBBA-CNR, prepared a questionnaire in order to address the legal, financial and administrative framework as well as evaluation of acceptability of the proposed measures by the sector with the final aim to identify the main barriers to the pick-up and interest of the new analytical technology of botanical composition of compound animal feeds, i.e. the FEED-CODE method. The activity performed in the framework of T7.1 were coordinated closely with activity in T6.2 - Pilot project. In particular, 3 demonstration events were arranged and hosted by the SME-AG’s during the Feed-Code project:
• Sofia, Bulgaria: 13th of November 2015 – hosted by the Associations of meat processors in Bulgaria (AMPB)
• Brussels, Belgium: 30th of November 2015 – hosted by Conseil Europeen des Jeunes Agriculteurs AISBL (CEJA)
• Rome, Italy: 26th of November 2016 – hosted by Consorzio Del Formaggio Parmigiano Reggiano (CPR) and Agricoltura è Vita Associazione (AèV)

As part of these demonstration events, the questionnaire was handed out to the participants and then collected by the end of or after the meetings. Totally 88 questionnaires was distributed to participants in these 3 events and totally 42 was collected, i.e. a response rate of 48%.
In addition, the questionnaire was made available as a web based survey to the members of CEJA, CPR and AèV. Totally, 15 respondents provided information in the web based survey.
The filled in questionnaires from the 3 demo events were returned to TI who has analyzed the data together with the results from the web based survey (D7.1).
The major results from end-user survey include:
• The majority finds it quite useful or very useful to control/analyze compound feeds.
• The majority finds it of high importance or medium importance to know the exact percentage of the compound feeds.
• The majority specifies that the Feed-Code analytical method is quite useful or very useful for them.
• The majority specifies that the Feed-Code analytical methods is important to them with regards to verify the composition of acquired feeds or to verify the purity of raw primary materials.
• The majority specifies that the major advantages of the Feed-Code methods is documentation of quality and fair trade.
• The majority specifies that major inconvenience is that analyzes are not require by law and high costs of services.
• The majority specifies that they to a high or medium extent will use the Feed-Code analytical method if this is provided as a service.
• The majority finds that it to a high or medium extent is of importance that the Feed-Code methods should be accredited according to international standards for laboratory analyses.
• The majority finds the cost of Euro 80 for analytical services to be too high or a little too high.
• The majority specifies that they will purchase services/send samples for analyses a few times per year.
• The majority specifies that they will send less than 50 samples for analyses per year.

Moreover, in accordance with point 15 of the list of objectives (revised D6.1) TI has also planned and arranged for benchmarking and validation of the new analytical methods developed in the FEED-CODE project following the provisions and requirements in ISO 17025: :2005: “General requirements for the competence of testing and calibration of laboratories”.
Originally, benchmarking test was supposed to be supported by the development and availability of the AOX system as a gold standard method but this was hindered by the difficulties and the delays UEVORA met in the preparation of reliable and consistent AOX-derived end-point PCR probes as well as the lack of several, reliable Taqman probes. It was then decided to replace this benchmarking approach by another one based on inter-laboratory comparison (ILC) that is comparing, on commonly shared feed samples, the data of the FEED-CODE method with existing, currently used feed analytical procedures. It turned out that the only ongoing system for feed qualitative and quantitative analysis is that based on the optical microscope (OM), and that no standards for feed measurements composition were/are available at European level.
As already mentioned in paragraph this led us to prepare our own reference mixtures designed to contain, in a common background of 12 different species (4 of which wheat, maize, barley and soybean) commonly abundant in feed for cattle, different and increasing amount, from 1 to 12% of the low abundant (sugar beet, sunflower, flax) or undesired species (cotton, peanut, rice and rapeseed). In this way we could provide, thanks to the work of internal validation, consistent quantitative data in the range of the 1-10% w/w presence of the minor or undesired ingredients.
Indeed, we run two different inter-laboratory comparisons (ILC), one organized in Italy and mastered by IBBA-CNR, the other run at the European level under the mastering of TI with the involvement of EUROFINS, a notorious analytical platform, as a third party to make the comparison fully trustable.

Refer to the Section of pdf file for a more detailed description of result n°3: FEED-CODE certification procedure and service, and related figures.
Potential Impact:
Route for the exploitation and future commercialization
The FEED-CODE market proposition is to “provide specific products (Kits) and certification services for the analysis of seeds or raw material, and more in general compound animal feed in order to verify the claiming and the actual composition declared in the label”.
To this scope, the FEED-CODE solution package aims at integrating the demand for innovative technical solutions for feed analysis and the services related to certification systems to quality and safety for feed.
By providing an integrated solution for European farmers (milk producers, meat producers, feed producers) and related associations, the FEED-CODE project has the ambition to cover user needs from a twofold perspective:
• the effective and cheap measurement of the exact composition of compound feed for food animal in order to assure frequent controls and in compliance with the new European standards;
• the food product quality and safety certification services.

In order to match the technology offer made available by FEED-CODE project and the users’ needs of the target groups identified, the partners defined and assessed a specific set of services and products with the aim to maximize the benefits and technological advantages achievable through the adoption of the combinatorial approach proposed.
FEED-CODE certification services (QUALITATIVE approach)
Application: to analyze and to certificate the qualitative composition of compound feeds and the purity of raw material
Users: producers of compound feed, private laboratories or control authorities
Value Proposition: FEED-CODE certification procedure allows to have information, record and traceability about compound feed composition, to be provided on the label or on request to the purchaser.
Estimated market price: about € 80 per single feed sample

FEED-CODE specific kits (QUANTITATIVE approach)
Application: to identify the presence (or absence) of a specific set of species in a compound feed.
Users: high quality (or PDO) products, where specific disciplinary of productions exist.
Value Proposition: FEED-CODE kit offers an easy, economic and fast way to provide high accuracy results, even traces, about specific target species content.
First user case: set of species for feed analysis of Parmigiano Reggiano cheese
Estimated market price: about € 100 per single sample

FEED-CODE business model
The initial exploitation strategy foresees the establishment of a FEED-CODE cauterized entity (such as NewCo or any other form of joint venture) ruled by the definition of a Joint Exploitation Agreement (refer to the D8.5) implying:
• the direct participation of the key technological and commercial partners of FEED-CODE Consortium;
• the establishment of preferential agreements with the Associations representing farmers (clients of the solution).
According to this strategy, the NewCo will act as a single contact point towards licensing the FEED-CODE process and methodology, as well as commercializing (or licensing) the FEED-CODE kit and carrying out certification service on demand. In fact, the NewCo could act both as product manufacturer (producing kits on demand for analysis laboratories) and services provider for farmers (realizing analysis for both certification and quantitative presence/absence of specific species).
This business model for exploiting the FEED-CODE application solution, mainly consists of a service to European farmers (milk producers, meat producers, feed producers), and/or a kit product for external laboratories or other control organizations/authorities, through the direct provision or the definition of commercial agreements with service companies:
According to the mission organization and main capabilities of each partner (refer to the details description of FEED-CODE Consortium partners reported in the D8.5) the preliminary assumptions considered for the definition of the roles into the NewCo are:
• technology and services’ provider: CNR, UEVORA, LABOR and TI
• commercial partner: FAI and MB
• lobbying/promotional partner: AèV and CEJA
• key clients (preferential agreements): CPR, RMA and AMPB

In this view, the Associations will be mainly involved in the promotion of the new labeling standard towards regulatory bodies, in order to push the commercial exploitation of the FEED-CODE results.
Moreover, consultation with stakeholders and decision makers has allowed to evaluate the optimal choice in terms of application model of the technology, to grant a bilateral cooperation between providers and buyers/users of compound feed and, therefore, to provide the most balanced and acceptable option.
However, based on the result of a preliminary stakeholders’ analysis carried out with reference to the actual Consortium composition, the partners agreed on the continue considering and evaluating an alternative scenario for exploiting the project results, even through the definition of licensing agreements with external Party(es). This need was easily deduced by the missing or low existence of critical skills and competences internally to the Consortium, such as industrial and commercial ones.

To this scope, the Consortium partners identified an alternative exploitation approach with the aim to better assess the relevant entry market barriers identified:
✓ uncertainty on the actual market demand
✓ expressed users’ need to have recognized the FEED-CODE method as reference ones or new standard for animal feed analysis and labeling certification
✓ accreditation of the FEED-CODE services provider
✓ significant additional investments required to industrialize and to commercialize the FEED-CODE services.

Moreover, the different positions and level of interest for the future exploitation and commercialization of project results registered internally to the Consortium, implied the need to define and agree two different business models: the first one (model A) to be deployed in the short-term period (at least 1 year after the end of the project), and the second one (model B) to be implemented in the medium-term period.
In particular, the short-term model A will be deployed with the common aims:
➢ to valorize the work carried out and results achieved by keeping the overall Background (TBP method) and Foreground (FEED-CODE method) knowledge internally to the Consortium. In this view, the main purpose is to continue working in a cooperative way on the industrialization of FEED-CODE services and products created as well as the validation of results at a larger scale;
➢ to assure the proper provision of the FEED-CODE services analysis toward end-users already demanding (both internal and external to the Consortium) by exploiting the project assets and in order to continue assessing the actual market demand. To this regard, the main purpose is to validate the feasibility and the actual implementation of the long-term exploitation plan, including the related investment and timing required.

Dissemination and communication strategy
The project dissemination working team, mainly composed by all AGs and SMEs involved in the project, worked with professionals who are trusted opinion leaders and are influential in their fields. Both formal and informal networks were used to “spread the word” about FEED-CODE. This approach was essential to the success of the dissemination activities during its entire timeframe, which were focused on increasing awareness of FEED-CODE initiative, including its products and services, to the entire value chain of stakeholders identified. In addition, the predomination aim of the dissemination activities were to facilitate the uptake of the results from the SMEs and AG’s and more in general from the target segment from the market.
This purpose was mainly carried out by using various communication channels and materials, but also organizing face to face meeting and/or participating in international fairs and local workshops.
The dissemination activities were implemented by the entire Consortium, and mainly by the AGs and SMEs, which are deeply involved in the feed sector and directly engaged in the promotion of the project towards their associated members and networks.
Indeed, the main objective of the dissemination in FEED-CODE was to make the project a highly visible project and to seek publicity for its results, services and products, informing and educating the community about the details of the project and its benefits as well as engaging with and getting feedback from the target groups. To this end, the channels chosen for disseminating the project were mostly used to:
1. present the project and its research work, and
2. inform the potentially interested parties about the progress of the project, both during its course and after completion.

At the same time, intermediate objectives were drawn, this having the advantage of supporting the monitoring of the activities and of outlining a path that can be easily followed by the SMEs. In particular, the most important objectives were:
➢ Raise awareness of the FEED-CODE project, products and services
➢ Informing and educating the community about the details of the project and its benefits
➢ Engaging with and getting feedback from the target groups
➢ Promoting the FEED-CODE project, products and services

All the dissemination activities were designed to send a key message to the target audience(s). In determining the key message, its communication medium, and timing, consideration was given to identifying key stakeholder needs, namely who was the “receiving” individual/organization, what they needed to know about the FEED-CODE project, and how that information could be communicated clearly and concisely (before the recipient loses interest in the message).

“The FEED-CODE Project aims to provide stakeholders along the food chain with an innovative animal feed certification platform and procedure to guarantee the quality of meat and dairy products through an automatic and simple identification method.”

The FEED-CODE target groups
Two main types of key stakeholders were identified and targeted as the audience of the dissemination of the project during the project implementation. These were:
Primary Audiences:
• Farmers
• PDO cheese producers and meat producers, and their associations(end users)
• Compound feed manufacturers

Secondary Audiences:
• Planners/policy makers at European, National and regional level
• Regulatory bodies
• Schools
• European, National and Regional authorities engaged in support for the agro-food sector

To this regard, it is worth to highlight as an effective communication required that all material and contents were developed by taking into consideration the needs of the above listed target audiences. Moreover, the project’s partners acknowledged that the major stakeholder groups have significant internal diversity. The dissemination materials and media were therefore developed to address the diverse needs and infrastructure available to each audience.
Communication tools
The communication tools selected for use were those most likely to reach the target audience with regard to the dissemination strategy and principles identified for the project. In this view the Consortium devoted significant effort to: the design of a FEED-CODE project logo; the creation and maintenance of a project web site; the design of communication material (flyer and poster) and updated versions when necessary; the creation of a video animation called “the adventures of TIBIP”; action to protect the FEED-CODE project name and logo; the implementation of a demonstration video for FEED-CODE, and the design and implementation of a web training tool for FEED-CODE self-learning.
The activities of raising public participation and awareness was considered the basis of FEED-CODE project success. The communication, therefore, was mainly based on a relationship of trust between the initiator and the target of the message.
In additional to the preparatory actions listed above, several actions were realized with this aim:
▪ On-line actions:
✓ Updating of the FEED-CODE website periodically,
✓ E-mail and e-newsletters promoting the project, its products and services as well as recent project activities.
✓ Articles and press releases were published in order to spread the information to a broad audience.
▪ Off- line actions:
✓ Dissemination workshops were organized with end-users to present the project and achievements and share the project’s products and services.

A description of further communication tools is reported in the following section 4.1.5 whilst a detailed list of all the dissemination activities performed during the entire project is reported in the Table A2.

Refer to the Section 4.1.4 of pdf file for a more detailed description of project impact in terms of both exploitation and dissemination activities, and related figures.
List of Websites:
FEED-CODE web site represented one of the most important assets for the dissemination of the project during the second period. FEED-CODE has a specific domain registered at the following web address:
Acting as a means of common communication, the website allowed successful collaboration with the industries’ actors as well as the involvement of actors in keeping up to date with the latest project activities.
The news and events page of the FEED-CODE website was particularly active during the entire duration of project. This page displayed the main activities accompanied by visual material, usually in the form of photos. Only during the second period, 16 posts were posted on the news and events page. The majority of these posts illustrated land mark stages of the project as well as smaller events and activities.

The logo
The building of the project’s image started with the design of a distinctive logo which was created with the purpose to reinforce the identification of the key elements, distinctive of the project’s results. This logo has been included in the design and production of the FEED-CODE website, leaflet and poster as well as in all the public and private communication material produced by the Consortium.
Although not foreseen in the initial dissemination plan, during the second period, steps were taken to trademark both the FEED-CODE name and logo.

In order to increase the impact of the dissemination actions towards the well selected target groups, further communication tools have been designed and implemented by the Consortium:

Video Animation
The TiBiP video animation was made visible and disseminated through the homepage of the official project web site, at the following link: The video animation link led to a YouTube page, shown in the figure below.
The main purpose of FEED-CODE video animation was to inform the general public about the importance of the quality of feed that is given to food producing animals, such as dairy cows for the PGI cheese or beef cattle to produce the PDO Parma ham for example.
The main concept behind the creation of this video was to show that milk and meat producers and cheese makers can now rely on TiBiP for their certification and control activities, because it can analyse an animal’s diet in such a way that single ingredients can be univocally identified.
Specifically, in the video, TiBiP - the genetic detective - tries to find the answer to the question: “How can we ensure the quality of meat and dairy products?”. Thanks to his efforts and inquiries, it is confirmed that the quality of cheese and meat largely depends on their ingredients. In turn, these ingredients depend on how the animals have been fed.

Demonstration Session Video Animation
An animated demonstration video was also created during the second period to effectively illustrate how FEED-CODE works in practice by demonstrating each single step of the process from the arrival of samples to the final analysis results. The video particularly focuses on the laboratory analysis, how the two software systems work and the overall accuracy of the analysis.
The demonstration video was realized in October 2015, and was filmed in the laboratories of IBBA-CNR in Milan, Italy, and was prepared with the contribution of IBBA-CNR and Labor. The language of the video was English, and the prime target audience of the video was commercial partners, research centres, public bodies, and the scientific community. The video has a duration of 9 minutes and 57 seconds, and is available on the Labor YouTube channel at as well as on the FEED-CODE website home page via

FEED-CODE Web Training Tool
A web training tool was designed and created to enable the target audiences and stakeholders to self-learn about the FEED-CODE project and its main findings. The purpose of the web training tool was to provide an informative and educational tool, that is both flexible and customizable, that is intended for two different target groups:
1. consumers, farmers, breeders, technicians of consortia, and
2. technicians of laboratories, feed manufacturers, operators of ports and customs.
The web training tool was created on a web learning environment owned by AèV, and consists of two different modules composed of learning units, available in Italian only at this stage. As a multimedia communication system, there are two separate training modules:
• A training module, called FEED-CODE, for the general public, policy makers, final users (for example the consortia and farmers) etc. which provides a general overview of the reasons for the establishment of FEED-COE and its expected outcomes, and
• A training module, called FEED-CODE Pro, for lab analysis professionals, operators of control agencies, feed producers etc., which provides details of the FEED-CODE procedure, instrumentation, and outcomes of the analysis in a highly detailed manner.
At the conclusion of each module, there is a self-assessment test, so that users may self-check their level of understanding of the topics covered. While AèV has coordinated the preparation of the storyboard and the implementation of platform, CPR, IBBA-CNR and Labor, were all responsible for the contents of specific units in both the modules. The web-training tool is available via

Refer to the Section 4.1.5 of pdf file for a more detailed description of project web site as well as other main communication tools, and related figures.