MycoKey develops new approaches to mycotoxin management: integration of output arising from the global knowledge harmonization, by involving international experts in gathering scientific knowledge on key topics, and obtained research data allows new smart and user-friendly MycoKey app. This system represents a step forward for scientific practical advises to growers, stakeholders and researchers.
The project is enhancing the EU-China dialogue, by involving the Chinese Consortium in linked research activities conducted at regional levels, and contributes to strengthen skills of young researchers and awareness about mycotoxins risks on human and animal health.
Alliances/scientific agreements with EU funded projects, especially with the MyToolBox project, are enhancing the international cooperation.
On-site applicable and ICT new integrated solutions are developed in the field of toxigenic fungi and mycotoxin monitoring along the chain. Integrated prediction models are under development for efficient crop management and stakeholders use. The approach is based on: a) risk models combination and control systems on site, b) development of new (multi)-mycotoxin screening methods on-site for multi-mycotoxin in situ and on-line analyses detection technologies for customized use in an open source environment.New validated detection methodologies and advances in knowledge on mycotoxins modified forms, which are of interest for legislators will be available.
In the field of prevention and intervention, the project is enhancing the potential of aflatoxins biocontrol in maize and peanuts, by applying it to emerging high risk areas in Europe (Italy, Serbia, Romania) and China. Sustainable control strategies based on natural compounds and biological control agents to be used in FHB and aflatoxin prevention are in progress and studied for market application.New less susceptible wheat genetic material is under selection. Less toxic and more effective new fungicides are tested. Novel post-harvest green solutions, sanitation techniques and cleaning equipment to reduce mycotoxins in food products are in progress for industries. New feed additives will be available, based on multi-mycotoxin detoxifying agents, and criteria and protocols validated and standardized, in line with EU regulations and EFSA guidelines. The safe use options solutions will release effective biological methods for mycotoxin detoxification and cost effective production of biofuel from mycotoxin contaminated biomasses.
Finally, 21 manuscripts were published in peer reviewed journals.
MycoKey solutions for the food and feed chain management – 3 years results
MycoKey is playing a significant role in the context of solutions to tackle biological contamination of maize, wheat and barley in the food and feed chain, by applying an integrated approach to reduce the risks associated to toxigenic fungi and related mycotoxins, specifically aflatoxins, deoxynivalenol, zearalenone, ochratoxin A and fumonisins.
The 31 project partners are generating innovative and integrated solutions to support stakeholders in an effective and sustainable mycotoxin management along food and feed chains, to be applied both in Europe and in China. The MycoKey actions focus on three areas: a) global knowledge and networking, in particular with China, input for regulations; b) innovative practical tools for farmers, end users and stakeholders for mycotoxin monitoring and management, c) reliable solutions, sustainable compounds/green technologies in prevention, intervention and remediation. The developed methodologies and tools are targeted for cost-effective application in the field and during storage, processing and transportation, including alternative and safe ways to use contaminated batches.
With regard to global knowledge and networking, a series of round tables gathering international experts allowed to update scientific knowledge on the main topics of mycotoxin contamination: biodiversity, genomics as well as metabolomics of toxigenic fungi; advanced detection methods for mycotoxins, predictive modelling for toxigenic fungi and mycotoxins; emerging and modified mycotoxins, toxicity and regulation. This MycoKey global and multi-disciplinary scientific network has been involved in working groups on specific crops: maize, wheat & barley, grapes, nuts & dried fruits. In addition, the project is turning the scientific knowledge and tools developed during the first 36M of MycoKey into easy to understand messages for end users and stakeholders. To this scope, guidelines for mitigation of mycotoxin contamination in maize and wheat, lay summaries and papers to give scientific support and suggestions included in the guidelines, were prepared. The MycoKey Charter has been launched as a supporting tool to gather information and involve policy makers and stakeholders to define common actions to tackle mycotoxin contamination at the global level. These actions were conducted both in Ghent and Wuhan and will continue in Belfast in 2019.
The project is contributing to enhance the dialogue with China, where the parallel integrated research plan is being carried out by 11 Chinese partners and funded by MOST- Ministry of Science and Technology, thus going to achieve the objective of contributions to reduce the frequent and severe mycotoxin contaminations in crops.
A strong commitment by the project Coordinator and EU partners fed and boosted the process of cooperation with China , through participation at main institutional events and meetings. Chinese partners play relevant roles in all Work Packages and project’s boards; they participated at the MycoKey training course in Europe, organised the successful 2nd MycoKey International Conference in Wuhan, hosted the Executive Committee meeting, the General Assembly as well as the round tables in 2018. They will organise a joint training course on food safety and an exploitation action with the Buhler group in 2019. Ten joint papers have been published so far, derived from different work packages on biodiversity of toxigenic fungi, mycotoxin detection and toxicology.
With regard to innovative practical tools for farmers, end users and stakeholders for mycotoxin monitoring and management, the MycoKey app represents a new integrated information system able to deliver information, links to tools and results/methods arising from the project activities to mitigate mycotoxin contamination. The MycoKey app was developed and integrated in the Akkerweb platform generated by Wageningen and partners. The app was presented and demonstrated during 7 international oriented workshops. The MycoKey app for cereals and maize will work globally and provides real-time and location-specific information on mycotoxin risks applying available mycotoxin risk models Both apps are currently free of charge and valorize weather data, satellite, sensor and science-based information for growers and other stakeholders for mitigation of mycotoxin risks. We are still working on the integration of Good Agricultural Practices (GAP) and customised advices. The tool will be used as a decision-making resource by stakeholders (e.g. growers, industry and food processers) to apply recommendations along the food and feed value chains.
In the field of mycotoxin monitoring, MycoKey developed rapid, reliable and validated detection tools (kits) for toxigenic fungi and multi-mycotoxins for on-site application of the main chain/toxin combinations: Maize – AFLA, FUM, DON, ZEA; Wheat/Barley – DON, ZEA, OTA. In addition, analytical tools (kits) were developed for traditional and alternative sampling methods (grains and dust).
Regarding toxigenic fungi and mycotoxin risks, a new model for AFLA and FUM joint prediction in maize is now available, which will support maize chain operators in mitigation; an updated model for DSS on fungicide application to reduce DON in wheat is available for farmers (expected reduction of DON by 20%). Furthermore, an on-site living sensor application tool was developed for famers: fieldmates are currently operational in BE, NL and IT for wheat and maize, but can be extended to any field grown crop. The weather data are integrated via network of dBases to an IT-dashboard. A rapid LAMP assay for in-field detection of toxin producers for wheat and maize is available.
In addition, new rapid multi-toxin test kits are availableand were validated to improve the process of management in food industries for food safety controls. These include a 3-myco DON/ZEA/FUM multiplex dipstick for maize, wheat and barley; a one-step multiplex strip test for AFB1/OTA/ZEA in maize, rice and peanuts; an aptameric multi-mycotoxin strip test; two fluorescence polarization immunoassay-based method for the simultaneous determination of DON and modified forms, for T2/HT2 and modified formsin wheat; and a real-time electrochemical profiling system for AFB1 in wheat and dried figs.
Four on-site dust sampling studies using the Rapidust® sampler were performed (two formaize and two for wheat). An LC-MS/MS method to provide data on multi-mycotoxin contamination in dust (and correlation with their content in maize and wheat) was developed, validated and applied to the analysis of maize, wheat and dust samples.
An assembly of novel toxicokinetic data for regulations on modified forms of DON, ZEA and AFB1 is currently finalised to improve process management of maize, wheat and barley. These datawill contribute to legislation and standard settings and a report for EFSA will be available by the end of 2019.
Reliable solutions, sustainable compounds/green technologies in prevention, intervention and remediation complete the contribution of the MycoKey project for the food safety.
In the field of mycotoxin prevention, the MycoKey solutions improve the breeding of less susceptible varieties/hybrids and the biological control of toxigenic fungi.
Highly tolerant varieties and hybrids were developed for breeding and field test. The identified commercial varieties and hybrids are promising with up to 70% DON reduction in soft wheat and,depending on the variety,with 30 to 60% DON reduction for durum wheat, while 13 promising maize hybrids with low levels of FER susceptibility and satisfactory grain yield resulted in30 to 80% FUM reduction.
Regarding the use of biocontrol agents/ natural compounds/ resistance inducers as well as cropping measures, new insights were derivedfrom the international research. For example, AflaSafe II is a new line of a successfully marketed biopesticide assuring between98% and100% AFLA reduction in maize. Furthermore, in Switzerland, a suitable inoculum production for the fungal antagonistClonostachys roseawas successfully developed , resulting in 60% DON and 30% ZEA reduction in wheat.
To improve field management practices, several new biological control agents have been developed for Fusarium head blight (FHB) control in wheat. These include (local C. roseastrains(DON reduction between69-85%);Bacillus velezensis RC 218 (DON reduction between 69 and 85% );Streptomyces spp.(DON reduction between50 and 75%); andPseudomonas piscium ZJU60 (biocontrol efficacy of 79%). In fact,Pseudomonas piscium ZJU60 andPantoea agglomeras ZJU23 were comparable with chemical fungicides phenamacril or tebuconazole, respectively (biocontrol efficacy of 80%). In addition,mustard-based botanicalsfor control of F. graminearum in wheat showed a DON reduction from 75 to 100%.
In the field of intervention, innovative methods have been developed, such as novel fungicide compounds effective against FHB, allowing DON reduction of 90% in wheat, introducing a new generation fungicides for FHB control, as well as new EOW treatment in field showing 80-100% OTA production in grape.
In the field of remediation, sustainable technologies for sanitation of contaminated crops post-harvest are available. They include:
- Ozone generation against Aspergillus flavus and Penicillium nordicum growth during storage, leading to 100 % inhibition of flavusand P. nordicum development in grains in wheat and barley;
- New natural chemical device for the control of flavus and P. verrucosum in cereals during storage, with 72% OTA reduction inwheat and 92% inbarley;
- Atmospheric Plasma Jet treatment on flavus in wheat, allowing a reduction of the fungal growth between 12 and 44%;
- UV treatments for the reduction of the flavus and P. verrucosum growth during storage, allowing significative reduction of toxigenic species(P. verrucosum) in green coffee, A. flavus in maize and walnuts.
- At industrial scale level, the combination of post-harvest cleaning technologies (mechanical and optical sorting) to be used by feed/food cereal processing industries significantly reduce contamination of mycotoxins in maize (AFLA: 60-90%, FUM: 65%, DON: 55%, ZEA: 100%).
New feed additives (multi-mycotoxins detoxifying agents) were developed to be potentially used by professionals in animal production and processing of animal food products; feed mills and feed additive producers. These feed additiveswere derived from two developed production processes, aimed to improve the efficacy of a yeast-based product in adsorbing ZEA (> 85%) , OTA and FB1 (>65%) (implementation and scale-up at industrial level), and to produce a newsurface modified bio-organoclay aimed to reduce the in vivo bioavailabiliy of AFB1, FB1, OTA, (> 90%) , andZEA (> 70%) level in feed (maize and wheat).The in vitro efficacy of the developed multi-mycotoxin detoxifying agents was confirmed by toxico-kinetic studies with rats and pigs by using the biomarker approach.
Microbial and enzymatic detoxification. New enzymes/microorganisms as safe use optionswere developed:
- Bacterial strains for reduction of OTA in wine (41-60%)and DON in wheat grains (100%);
- Purified recombinant proteins / cloned bacteria (DON reduction: 50-70%) and Laccase enzyme purified from Pleurotus (reduction of AFLA: 90%, ZEA: 100%, FB1: 40%, T-2: 40%)
Mushroom strains(Pleurotus eryngii, Agaricus bisporum, Lentinula edodes,Cyclocybe aegerita, Hypsizygus tessellates -Shimeij), (reduction of AFLA: 65-100 % of ; FB1: 67-100%, ZEA 91-100%)
- biogas production was not affected by mycotoxin contaminated grains as feeding substrate, and aflatoxins and FB1 were reduced by 90%. Bioethanol was efficiently produced in bioreactors with selected tolerant yeast strains from mycotoxin contaminated biomasses with good yields and productivities.
Work performed and main results achieved in 18 Months since the beginning of the project in the following Work Packages:
GLOBAL MYCOTOXIN KNOWLEDGE
- review of scientific updates, adapting existing knowledge on mycotoxins for stakeholders’ use;
- a predictive model transfer (AFLA_maize) and guidelines for minimization of mycotoxin contamination ready;
- two Round Tables organized in Ghent and the MycoKey Charter web-site launched.
TOXIGENIC FUNGI MONITORING
- AFLA&FUM predictive modelling data acquirement available;
- sequence data for public on-line Databases for fungal species identification;
- On site LAMP assay developed. Whole genome sequencing, information on new genotypes and distribution of mycotoxin gene clusters;
- Use of environmental on-site sensors.
- alternative practical method and device for grain dust sampling in maize under development;
- a multi-mycotoxin method to detect contamination in dust (and correlation with their content in maize) under validation;
- Multi-mycotoxin test kits for on-site screening (including UNISENSOR DON/ZEA/FUM multiplex dipstick test prototype) under development;
- Modified mycotoxins and their in vivo toxicokinetics research is ongoing.
PREVENTION IN THE FIELD
- biocontrol of Aspergillus flavus to prevent aflatoxin contamination in maize (aflatoxin reduction range: 98%-100%);
- selected intercropping systems reduced Fusarium Head Blight (FHB) disease;
- identification and formulation of biocontrol agents allowing reduction of FHB infected wheat spikelets and DON contamination;
- selection of less susceptible maize hybrids to Fusarium Ear Rot and wheat varieties to FHB.
- potential biocontrol agents identified to prevent FHB in cereals;
- new fungicides tested in Argentina, Belgium, China and Italy;
- the efficacy of electrolyzed oxidizing water on OTA contamination in wine proven;
- laboratory scale stand for ozone treatment manufactured;
- non-thermal plasma system established for wheat;
- identification of potential technologies for malting industry.
- proven efficacy study of industrial-scale cleaning equipment in reducing aflatoxin content in naturally contaminated maize (range of aflatoxin reduction: 60%-90%);
- In vitro tests for binding simultaneously mycotoxins (to be tested in vivo);
- Study of microbial proteases efficacy for OTA degradation in wine processing;
- Proven efficacy of industrial mushroom Pleurotus eryngii in AFB1 degradation (up to 100%) in vitro and cultivation substrate;
- Pleurotus laccase enzymes proven to be effective for in vitro degradation (up to 100 %) of AFB1, ZEA, AFM1.
ICT SOLUTIONS FOR CHAIN MANAGEMENT
Interface of the MycoKey app designed, beta version developed. The companion app OnTheSpot was generated to allow georeferenced recording of events and remarks in the field using a mobile phone.
COMMUNICATION, DISSEMINATION, EXPLOITATION
- presentation at international conferences and meetings;
- the 1st MycoKey Conference was successful held in Ghent, in September 2017;
- 3 Short learning visits for young students are in progress;
- 2 training courses have been organized in Italy in 2017.