ASTER is organised in 6 Work Packages (WP): coordination and management (WP1), characterization and enhancement of belowground biodiversity (WP2), characterization and enhancement of aboveground biodiversity (WP3), natural derived compounds (WP4), implementation of sustainable management methods (WP5), uptake, dissemination and formation (WP6). All WP are managed by project coordinator and a WP leader. When needed the WP is divided into specific tasks (each referring to a task leader).

Leader: Emilio Guerrieri

Objectives: to guarantee the overall scientific and administrative coordination, management and success of the project
Description of work:
All administrative and coordination activities fall within this WP including:

– Organization of meetings: the coordinator will maintain regular communication with the partners, WP and task leaders via email, phone, skype; two meetings per year will be organized (including kick-off and final meeting) to present and discuss the results and the advancement of the works and to present/remove unforeseen obstacles to scheduled activities and deliverables for which actions will be taken. Administrative and financial issues will be discussed, together with intellectual property and ethical aspects.

– Preparation of reports: the coordinator will: provide guidelines and set deadlines for the presentation of financial and scientific reports; monitor the respect of the schedule, with the help of the unit and WP leaders; maintain regular communication with the PRIMA Foundation.

– Constitution and management of Stakeholder Interactive Platform (SIP): meant to ensure a constant flow of information between academic and non-academic partner/supporters and to aid in planning field experiments (see support letters).

– Data management plan and protocols: data will be treated as confidential and shared on web/cloud systems accessible with passwords; Common Protocols for Research Activities (CPRA) will be discussed and approved at the kick off meeting; the laboratories involved in ASTER will follow the rules for Good Laboratory Practice (GLP).

Deliverables (brief description and month of delivery)

D1.1 Common Protocols for Research Activities (CPRA) (M2)
D1.2 Stakeholder Interactive Platform (SIP)(M2)
D1.3 Minutes of Consortium Meetings (M1, M8, M14, M20, M28, M36)
D1.4 Financial Reports (M14, M26, M36+2)
D1.5 Scientific Reports (M14, M26, M36+2)

Leader: Sonia Labidi

Objectives: Characterization and enhancement of telluric microbial biodiversity to test and select the best strain/consortium for tomato growth, nutrition and protection against biotic and abiotic stresses.

Task 2.1 Characterization of soil microbial biodiversity (M3-M27)
Leader: Valeria Todeschini (P2). Participants: P2, P4, P11, P12, P13 P14, P15

Characterization of soil microbiota in the field targeting: PGPR (P2, P15), mycorrhizal fungi (P2,P11, P12, P14), antagonist fungi (Trichoderma)(P13), entomopathogens (Beauveria)(P13).

Methods: soil sampling (general) and plant trapping (Sorghum for mycorrhiza); isolation/extraction (plate cultures for bacteria); mass propagation; morphological characterization; biochemical characterization (P11, P13, P14, P15); bacteria and AMF molecular characterization by pyrosequencing using rRNA genes (16S, 28S, 18S) (P2, P4); taxonomic assignment to species level by blasting reads against a core set of the RDP3 database for bacteria (Wang et al. 2007. Appl Environ Microbiol 73: 5261-5267) and against a AMF specific database reported in Cesaro et al. (2020 – Mycorrhiza 30: 601-610; 2021 – Front Microbiol 12: 676610); bioinformatics analysis (P2); calculation of biodiversity indexes (P2); microbial selection according to the NGS analysis (P4). Bacterial strains potentially expressing plant growth-promoting traits (based on literature and species identity) will be characterized for indoleacetate production, phosphate solubilisation, siderophore production, nitrogen fixation and ACC deaminase activity (P2, P13).

Task 2.2 Testing the effect of beneficial microbes on tomato nutrition (M10-M28)
Leader: Naima El Ghachtouli (P15). Participants: P2, P5, P11, P12, P14

Laboratory and field tests (open, protected) run with selected commercial (first year) and/or native microbial species/strains belonging to Trichoderma (P11, P12, P14), Glomus (P11, P14), Bacillus (P15), Pseudomonas (P15) and tentative consortia to assess their effect on the following plant parameters linked to nutrition (P2, P11, P12): root and shoot biomass production; water content; yield; mineral up-take; photosynthetic pigments concentration (chlorophyll a, b and carotenoids) by spectrophotometer); photosynthetic performance in vivo on leaves previously dark-adapted for one hour by Handy PEA; photosynthetic activity in vivo by portable chlorophyll meter (SPAD); main micro- and macronutrients concentration in the different plant organs by (ICP-MS) after dry samples digestion in nitric acid.

Task 2.3 Testing the effect of beneficial microbes on tomato protection (M4-34)
Leader: Michelina Ruocco (P1). Participants: P4, P6, P8, P9, P11, P13, P14, P15

Laboratory and field tests (open, protected) run with selected and/or native microbial species/strains (if available from year 2) to record plant damage, plant resistance against (and recovery from) light and moderate drought stress (P11, P13, P14, P15)(seeVolpe et al. 2018. Front Pl Sci 9: 1480 for standard protocols), plant direct resistance to pests and pathogens; attraction in wind tunnel and olfactometer bioassay and performance of natural antagonists with the following combinations:

Partecipant number microbes targeted stress antagonists
1 Beauveria bassiana, ≥2
native strains (1 AMF, 1 root
symbiont), ≥1 consortium
aphids, Tuta absoluta Aphidius sp.,
4 commercial strains of
Trichoderma sp and Glomus
Tuta absoluta Dicyphus cerastii
6,8,13 ≥2 native strains
(Trichoderma, Glomus,
Bacillus, Pseudomonas,
Beauveria, Isaria), ≥2
Tuta absoluta, Bemisia
tabaci, Tetranychus
urticae, Aculops
Nesidiocosis tenuis,
phytoseiid mites.
9 commercial Trichoderma,
Glomus and Bacillus
Fusarium oxysporum f.
sp. lycopersici

Deliverables (brief description and month of delivery)

D2.1. List of main soil microbiota components (ex ante and ex post ASTER activities). (M12+M18)
D2.2. Biodiversity indexes in selected country areas. (M18)
D2.3. Selection of microbiota (strains, consortia) for tomato nutrition. (M34)
D2.4. Selection of microbiota (strains, consortia) for tomato protection. (M34)

Leader:WP leader: Alberto Urbaneja

Objectives: characterization of Hymenoptera as indicators of biodiversity in tomato crops; selection of companion plants to improve nutrition, protection and pollination in tomato crops; set up of protocols for the use of natural  antagonists

Task 3.1. Characterizing insect biodiversity targeting Hymenoptera (M1-34)
Leader: Massimo Giorgini (P1); Participants: P3, P4, P5, P6, P7, P8, P9, P10, P11, P12, P14

Each participant will select 4 open-tomato fields where a standard IPM program representative of each study area has been applying. Four collections/year are planned in early spring, summer,  idautumn, beginning of winter by: i) malaise traps (≥2/field to be adapted to field dimensions), short term samplings of 3 days/period; ii) UV-bright Pan Traps (white, yellow, blue) (1 trap set/3.000 mq), bee hotels (1/3000mq), transect walk and floral observation alongside the test crop and within it; iii) vacuum sampler (16 plants per field will be vacuumed, 10 second sucks/plant). Individuals will be determined by integrative characterization including morphological identification (at least at genus level) by available keys and comparing the material with that stored in main EU collections (e.g. Natural History Museum of London- which the coordinator is a scientific associate of) and molecular characterization targeting main taxonomic genes (COI, 28S). Characterizations relative to Year 1 will define the ex ante biodiversity indexes to be compared with those relative to Year 4 (ex post) in relation to the application of project results (see WP5).

Task 3.2. Testing the effect of companion plants on plant nutrition, protection and pollination. (M3-24)
Leader: Lucia-Zappalá (P3) Participants: P1, P4, P5, P6, P7, P8, P9 P10, P11, P12, P14

The tentative list of test plants as discussed with stakeholders will include: Tagetes sp., Phaceliatanacetifolia, Cnidium sp, lavender, lucerne, Fagopyrum esculentum, Plantago sp.,  Anethum graveolens, Sesamum indicum, Verbena sp, Lobularia maritima, Vicia faba, Pisum sativum and vetch. Initially in controlled semi-field / greenhouse conditions, the individual effect that each of the selected companion plants may have on the nutrition, protection and pollination of tomato plant will be studied in three separated experiments. The common experimental protocol includes: planting in the same pot (with sufficient soil volume for the normal growth of both plants) a companion plant and tomato (control pots consist of 2 tomato plants). i) Plant nutrition test (P6): recording plant height, root development, number of leaves, leaflets, flowers, fruits. ii) Plant protection test (P5): recording the development and infestation capacity of Tuta absoluta (P1, P4, P6, P8), Phaenacoccus solenopsis (P9),Tetranychus urticae and Bemisia tabaci (P6, P8), along with the infection capacity of the tomato yellow leaf curl virus (TYLCV) by infected populations of the whitefly (P6, P8). The influence of companion plants on the attractiveness of the antagonists Aphidius sp., Trichogramma achaeae and Cryptolaemus montrouzieri will be recorded in olfactometer and wind tunnel bioassay (P1). iii) Pollination test: recording the number of flowers set using bees as pollinators (P1, P3). From the results of this first phase (M3-12), ≥ 3 best companion plants that give the best results will be selected for field test. Six tomato open fields among the project consortia will be selected (Portugal-P4; Greece-P7; Turkey-P9; Tunisia- P10, P11; Algeria -P12; Morocco -P14). Field trials will be designed with 1: 1; 1:10 and 1:20 densisties of companion:tomato plants. In each experimental field, the same parameters as for controlled semi-field / greenhouse tests will be recorded.

Task 3.3. Testing natural antagonists against tomato pests and pathogens (M3-M24)
Leader: Alberto Urbaneja (P5) Participants: P1, P4, P6, P7, P8, P9, P10, P11, P12, P14

Combinations of antagonists used in tomato crops (inoculation of soil microbes, use of predator mirids, ladybeetles, entomopathogen nematodes and/or fungi) will be tested under three water tress
levels (normal, light and moderate stress). Main targets of these tests are Fusarium spp., Tuta  absoluta and Phaenacoccus solenopsis. Plots will be chosen in the same locations as Task 3.2.The strategies tested include the inoculation of soil microbes and the release of predator insects or entomopathogens in the nursery before transplanting versus inoculation of soil microbes in the nursery and release of predator insects or entomopathogens after transplanting. For each combination/treatment four replicates will be conducted (24 cages per field). Each replicate will be conducted in an insect cage (3 x 3 x 2m) in which 9 tomato plants will be planted. During year 1 commercial strains of beneficial soil microbes will be used whilst from year 2 tests will be run with a selection of native strains and consortia from WP2 (if available). Tested predatory mirids will be fed with Ephestia kuehniella weekly and their population dynamics recorded throughout the growing season. The species of predatory mirids will be selected for each area based on the previous natural occurrence of three main species (Nesidiocoris tenuis, Dicyphus cerastii and Macrolophus pygmaeus). In Turkey (P9), 3 different protocols for the release of Cryptolaemus montrouzieri in terms of timing, quantities and host densities, will be tested (P9) against the emerging infestation of mealybugs. In Italy (P1) preliminary greenhouses tests will assess the ability and the efficiency of bumblebees to carry and deliver antagonistic fungi. Contamination and delivery will be recorded on visited plants using coloured powders and commercial bumblebees colony boxes modified for the purpose (3 replicates; 100 bumblebee individuals each).

Deliverables (brief description and month of delivery)

D3.1. List of Hymenoptera morphospecies (ex ante and ex post ASTER activities) (M16, M35)
D3.2. Biodiversity indexes in selected country areas (M16, M35)
D3.3. List of companion plant (including wildflowers) selected for tomato nutrition (M24)
D3.4. List of companion plant (including wildflowers) selected for tomato protection (M24)
D3.5. List of companion plant (including wildflowers) selected for tomato pollination (M24)
D3.6. Species and use protocols for main antagonists of tomato pests and pathogens (M24)

WP4: Natural derived compounds
Leader: Farida Benzina

Objectives: Selecting tools for the sustainable protection/nutrition of tomato
Common protocols for research activities (CPRA) agreed during the kick-off meeting

Task 4.1 Testing the effect of essential oils (EO) and Hydrolates (H) on plant resistance (direct, indirect) to biotic and abiotic stresses (M3-30)
Leader: Antonio Biondi (P3) Participants: P1, P4, P5, P6, P7, P10, P12, P13

Laboratory and semifield tests to calculate the dose response curves of specific EO and hydrolates (H) of pathogens, pests and antagonists attractiveness (see task 3.2 for details) under different water regimes (light and moderate draught) with the following combinations:

Participant n. compounds targeted
1 EO and H from Chrysanthemum
coronarium and Achillea compacta
aphids, Tuta absoluta Aphidius sp.,
4 EO and H from Chrysanthemum
coronarium and Achillea compacta
6 EO from garlic, Ocimum basilicum Tuta absoluta,
Tetranychuns urticae
12 EO from garlic, lemon grass, olive
tree, Eucalyptus, onion, nettle
Fusarium oxysporum
13 EO and hydrolates from Chrysanthemum coronarium and
Achillea compacta
Fusarium oxysporum
f. sp. lycopersici,
Tuta absoluta

Task 4.2 Testing bioderived metabolites as elicitors of resistance (M10-28)
Leader: Apostolos Kapranas (P7). Participants: P1, P5, P12, P13.
Laboratory tests to calculate the dose response curves of different bioderived metabolites and application method on selected pests/pathogens mortality and on the attraction of natural antagonists with the following combinations:

Participant n. compounds targeted
1 VOC induced by Trichoderma and
Glomus colonization
Tuta absoluta Trichogramma
7 supernatant of EPN bacterial symbionts
of Steinernema carpocapsae, S. feltiae
and Heterorhabditis bacteriophora
Tetranychus urticae Phytoseiulus
sp., Neoseiulus
sodium alginate Fusarium oxysporum
f. sp. lycopersici;
Tuta absoluta, T.
urticae, whiteflies
12,13 lipopeptides (biofilms), phosphatases,
proteases, siderophores, HCN, ATB,
Enzymes, peptaibols, mycotoxins ,
polyketides, pyrones, terpenes,
metabolites derived from amino acids
and polypeptides
Fusarium oxysporum
f. sp. lycopersici,
Fusarium oxysporum
f. sp. albidinis and
Botrytis cinerea

Task 4.3 Production and testing in the laboratory and in the field the effect of crop residues on
plant nutrition and protection from soil pathogens (M5-M30)
Leader: Bouamri Rachid (P14), Participants: P10, P12, P13, P14.
Valorization and hygienization of tomato residues by co-composting and vermicomposting with olive industry waste (liquids, pomace) and/or manures; production of small-scale tea compost and vermicompost tea production; combining vermicompost, compost tea and vermicompost tea with biochar and arbuscular mycorrizal fungi (production of glomalin). Selected combinations will be tested in laboratory and small field plots for their effect of on soil chemical parameters (nitrogen, organic matter, phosphorous, potassium), plant nutrition (especially P), bioprotection against soil borne pathogens (Fusarium spp. and nematodes) (P10, P12, P13, P14). Crude olive pomace will be additionally tested as a substrate for culturing symbiotic microorganisms (Pseudomonas, Bacillus and Trichoderma) for the production of biopesticides by Fermentation in Solid Media (FMS) (P13).

Deliverables (brief description and month of delivery)

D4.1. List of EO and H and relative doses for tomato protection from main pests (M30)
D4.2. List of bioderived metabolites and doses as elicitors of tomato resistance (M28)
D4.3. List and protocol for the use of crop residues (M30)

Leader:George Broufas

Objectives: combining the selected tools and strategies into a holistic protocol for the management of tomato crop; to assess the overall economic, social and environmental impact of ASTER results.

Task 5.1. Combining tested tools into a holistic management protocol (M20-M35)
Leader: İsmail Karaca (P9); Participants: ALL.

Selected tools (soil microbes, companion plants, natural antagonists, natural derived compounds) and strategies developed in WP 2, 3 and 4 will be tested in Italy, Portugal, Spain, Greece, Turkey, Morocco, Algeria, Tunisia (≥2 fields/country) in a holistic management model and compared with local management protocols. Degree of reduction of chemical fertilization will be chosen depending on the soil fertility measured with nutrient content and other soil traits (including, but not limited to, pH, soil organic carbon and carbon exchange capacity) before starting the experiment. At least two plant species or mixtures will be established as companion plants in narrow strips within the  crop or at field margins. All other cultivation practices, if not clearly specified will follow the common management protocols of each country. Throughout the cultivation period, the experimental plots will be sampled at least every other week to: check for pest occurrence (Bemisia tabaci, Phaenacoccus solenopsis, Tetranychus urticae, Aculops lycopersici, Tuta absoluta.) and disease infestation index (for Fusarium sp., Verticillum sp., Phytophtora sp.); release the most appropriate biocontrol agents tested in WP2, 3 and 4 if needed; sample the natural population densities of biological control agents and pollinators. Biodiversity indices will be calculated as described in WP2 and WP3. Amount of culturable microbes soil microbiota (as in WP2) and fertility indices (including dissolved organic carbon and nitrogen, mineral nitrogen, phosphatase activities) will be measured before and after each experiment.

Task 5.2 Assessing ASTER impact on tomato harvest (M22-M35)
Leader: Valeria Todeschini (P2) Participants: P4, P5, P11, P14

The total fruit yield, weight, the average fruit number-per plant and per-treatments will be evaluated during the growing season in the experimental field. Fruit dry biomass and water percentage from fresh fruit homogenate dried at 60 °C for 7 days will be calculated. Chemical quantification of main tomato parameters (pH, acidity, lycopene and b-carotene concentration by HPLC) will be determined on tomato fruit homogenate obtained from pooled fruits (from each treatments) previously frozen in liquid nitrogen, blended and stored at –80°C (P2, P4, P5, P11, P14). The fruit firmness will be evaluated with the use of a bench top penetrometer on 10 fresh collected ripe fruits. All parameters recorded will be compared with those realized following conventional management protocols.

Task 5.3 Assessment of economic, social and environmental impact of ASTER (M1-M36)
Leader: Ali Abidar (P14), Participants: P3, P9

The economic, social and environmental impact of ASTER will be assessed by meetings, questionnaires and focus groups (via SIP) to gather information on the main features of actual productive chain in selected partners’ countries. An analysis of the elements motivating the adoption of the production methods by producers will be conducted via SIP to evaluate their potential interest (P9, P14). Life Cycle Assessment (LCA) and a cost-benefit analysis will be performed (P3, P14). The Life Cycle Assessment (LCA) will assess the trade-offs between the use of inputs and the production techniques used in order to identify the most sustainable ones. The cost-benefit analysis (CBA) will be used to estimate the costs and benefits of decisions in order to find the most costeffective alternative considering: Direct costs, Indirect costs/Intangible costs, Direct benefits, Indirect Benefits. These aspects, if properly used, can also be a plus in terms of marketing policies by the players in the supply chain (P3). Results may be expressed by internal rate of return, net present value and benefit-cost ratio. The project will lead to new products for different markets:
direct food consumption, food transformation, forage for livestock, cosmetic and pharmaceutical industries. Their potential for these different markets will be assessed. A survey of potential users will be conducted for this purpose (P3, P14).

Deliverables (brief description and month of delivery)

D5.1. A user friendly, protocol for agroecology management of tomato crop in small farms (M35).
D5.2. Correlation analysis between management protocol and harvest (quality, quantity)(M35).
D5.3. Cost-benefit analysis of crop production protocol developed by the ASTER consortium (M35).
D5.4. Life Cycle Assessment of the new tomato production protocol (Report: R) (M35).

Leader: José Carlos Franco

Objectives: Generate ASTER impact through active dissemination, communication and formation

Task 6.1 Communication – Leader: Pasquale Cascone (P1). Participants: ALL
Project aim and start will be widely advertised to local press/TV, schools and universities, governmental institutions (e.g. the Wilaya farming chamber, NPPOs) in all participating countries by interviews (at least 1 interview per participant; total > 15), explicative leaflets and posters (at least 1 leaflet/poster per participant; total > 15). Lectures will be offered in primary/secondary schools and Universities on the themes of agroecology, sustainability, safe food, climatic change and on how ASTER addresses them (at least 2 per participant total > 15 x 2 = 30).

Task 6.2 Dissemination- Leader: Lucia Zappalà (P3); Participants: ALL
ASTER model will be promoted and disseminated starting from SIP whose participating  stakeholders guaranteed their contribution. Project page on the web with differential levels of access (restricted to project parteners, public) and main social media will be created in English, French, Arab (including local varieties from the project partners), Italian, Spanish and Portuguese and updated in real time (and at least every 3 months for public) to disseminate the progress of research activities and results. Links to project webpage will be created on the Institutional pages of project partners. In selected countries, open workshops (≥2/country) will be organised with web connection available to reach the widest possible audience. Scientific publications (≥10 Gold open access) for high impact and local journals and congress presentations (≥ 6) are planned. Demonstrating videos (≥2 including presentation and main achievements of ASTER) will be realized in the field with subtitles/doubling in languages of participating countries.

Task 6.3 Formation – Leader: Rachid Bouamri (P14); Participants: P1, P7, P10, P11, P12, P13
Activities organized to promote the use of agro-ecological practices elaborated by ASTER include: (i) 3 on-site training courses targeted to farmers in Mediterranean-producing Countries in Morocco, Algeria and Tunisia with modules of 15 people each and lasting 3 days (at least 1 module/year in selected Countries), (ii) installation of 2 permanent didactic farms, in Greece (P7) and Morocco (P14). All training courses will be video-recorded and subtitled in languages of participating countries.

Deliverables (brief description and month of delivery)

D6.1. Pages on Web and social media (M2)
D6.2. WorkShops and webinars (WS M14, M28)
D6.3. Communication and divulgative material (from M3)
D6.4. Lectures (M6, M12, M18, M24, M30, M35)
D6.5. Training courses (+ videos) (from M24)
D6.6. Publications (from M26)
D6.7. Didactic farms (from M24)

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