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Phenotyping for Resilient Fruit Systems Production

Implementing experimental designs and phenotyping methods to select local ecotypes or varieties with specific traits, including tolerance/resistance to biotic and abiotic stresses, horticultural and fruit quality traits. Objectives: Apply established phenotyping protocols, characterize phenotypic variability, identify genotypes for pre-breeding, analyze genotype-environment interactions, and produce phenotypic data for genetic analyses.

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Phenotyping for Resilient Fruit Systems Production

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  1. WP1 - Phenotyping for traits relevant to resilient fruit systems production Co-Leaders: Rania Ahmed (ARC) Iban Eduardo (CRAG) FREECLIMB PRIMA Meknes 2019

  2. WP1 - Phenotyping for traits relevant to resilient fruit systems production Objectives Aim of WP1 is the implementation of experimental designs, phenotyping methods and tools to improve screening/selection of local ecotypes or varieties for traits defined by already set ideotypes (see section 1.3 a). Where possible we will take advantage of already validated protocols and transfer knowledge between species on abiotic and biotic stresses, while also monitoring horticultural and fruit quality traits. Specific objectives of WP1 are to: ● Apply already established phenotyping protocols for assessing tolerance/resistance to biotic and abiotic stresses, suitable for field and/or controlled conditions ● Characterize phenotypic variability available within MB germplasm banks ● Identify genotypes to be used in pre-breeding for MB conditions ● Analyze GxExM interactions for extrapolating ideotype performance under different conditions ● Produce phenotypic data for genetic analyses (WP2) (materials shown with number and asterisk [#*]; further information in FREECLIMB plant material Table 2 FREECLIMB PRIMA Meknes 2019

  3. WP1 - Phenotyping for traits relevant to resilient fruit systems production Fruitcropsideotypes: Stone fruitsRootsock: adaptation to sub-optimalwatersupply and differentsoiltype (in particular calcareoussoils); nitrogen use efficiency; root-knotnematoderesistance; Almond: Scion: resistance/tolerance to late frostdamage; Sharka (PlumPox Virus, PPV) resistance Apricot: Scion: resistance/tolerance to late frostdamage; low-chillingrequirements; resistance to blossombrownrot (Moniliniaspp.); PPV resistance Peach: Scion: low-chillingrequirements; highwater use efficiency; resistance to blossom and fruitbrownrot (Moniliniaspp.), powderymildew, leafcurl and PPV. Olive. Scion: tolerance/resistance to olive fruitfly (B. oleae); earlyflowering to avoidheat-stress; adaptation to high air temperatureduringflowering; Rootstock (orself-rooted): resistance to Verticilliumwilt, soilsalinity and sub-optimalwateravailability. Citrus. Scion: tolerance to ‘Mal secco’. Rootstock: resistance/tolerance to Phytophthora, soilsalinity, calcareoussoils and sub-optimalsoilwateravailability. Grape. Scion: downy and powderymildewresistance. Rootstock: Grape phylloxera (D. vitifoliae, Fitch 1855) resistance, adaptation to a range of soiltype and wateravailability. FREECLIMB PRIMA Meknes 2019

  4. WP1 - Phenotyping for traits relevant to resilient fruit systems production Task (T) 1.1 Evaluation of plant response to biotic and abioticstresses in germplasmcollections. M 1-24. Task leader: INRA-GAFL. Partnerinvolved: UMIL, INRA-BFP, CRAG, IOSV, CREA, ARC, CU, INRA-MO, IO, UFMC, ENSA. T1.2 Phenotypingsegregating progenies to deciphergeneticdeterminismfortraitsrelevant to predefinedideotypes. M 1-24. Task Leader: CU. Partnersinvolved: UMIL, CREA, CRAG, INRA-GAFL, INRA-BFP, CU, IO. T1.3 Environment and managementeffectonadaptivetraits. M 1-24. Task Leader: IPB&GR. Partnersinvolved: UMIL, CRAG, IOSV, IO, INRA-MO, ENSA). T1.4 Omicsanalysis of environmentaladaptationmechanisms. M 12-36. Task Leader: CRAG; Partnersinvolved: UMIL, INRA-MO, ENSA, IPB&GR. FREECLIMB PRIMA Meknes 2019

  5. WP1 - Phenotyping for traits relevant to resilient fruit systems production Task (T) 1.1 Evaluation of plant response to biotic and abioticstresses in germplasmcollections. M 1-24. Task leader: INRA-GAFL. Partnerinvolved: UMIL, INRA-BFP, CRAG, IOSV, CREA, ARC, CU, INRA-MO, IO, UFMC, ENSA. • This task will implement protocols (including eco-physiological models) to run large-scale phenotyping on germplasm collections for traits related to environmental adaptation (abiotic stress and pest/disease tolerance/susceptibility); horticultural and fruit quality traits will also be assessed by routine methods. • To: • identify resilient parents to be used in further crosses to pyramid targeted traits (T3.1) • identify promising selections and/or putative varieties to be recommended for MB growing areas • collect data for GWAS studies (WP2). FREECLIMB PRIMA Meknes 2019

  6. WP1 - Phenotyping for traits relevant to resilient fruit systems production Task (T) 1.1 Evaluation of plant response to biotic and abioticstresses in germplasmcollections. M 1-24. Task leader: INRA-GAFL. Partnerinvolved: UMIL, INRA-BFP, CRAG, IOSV, CREA, ARC, CU, INRA-MO, IO, UFMC, ENSA. In almond: - collection of #50 accessions will be phenotyped for bud phenology, floral induction and anomalies, and for date of fall of leaves (marker to heat stress) (INRA-GAFL) [12*]. (150-180 accessions) In apricot: - a germplasm collection of wild accessions (Central Asia and Chinese natural populations) (INRA-BFP) scored for Sharka resistance will be phenotyped for bud phenology, floral induction and anomalies in two contrasted sites of INRA (INRA-GAFL) [10*] to be grafted and planted in Avignon. Traits: if lucky last year of the project, but probably not possible… Date of fall of leaves, Monilia on lower irrigation ; bacterial canker - 100 accessions including landraces, local and breeding cultivars (UMIL) already being phenotyped for Sharka resistance and brown rot resistance on flowers will be phenotyped for chilling/heat requirements. - a collection of local apricot varieties (including rootstocks) will be assessed for chilling requirement and drought tolerance (UFMC) [10*]. Chilling requirements will be also assessed in a collection of #34 local and commercial apricot cvs (IPB&GR). FREECLIMB PRIMA Meknes 2019

  7. WP1 - Phenotyping for traits relevant to resilient fruit systems production Task (T) 1.1 Evaluation of plant response to biotic and abioticstresses in germplasmcollections. M 1-24. Task leader: INRA-GAFL. Partnerinvolved: UMIL, INRA-BFP, CRAG, IOSV, CREA, ARC, CU, INRA-MO, IO, UFMC, ENSA. In citrus: - germplasm collections (including tetraploid lemons accessions) and selections will be tested for tolerance/susceptibility to Malsecco[1*] (CREA) - several rootstocks selections (diploid and tetraploid) will be tested for Phytophthora resistance as well as adaptability to saline water and drought stress [2*] (CREA, UFMC) In grape: - #100 V. vinifera accessions from the Caucasian region will be tested for resistance against powdery mildew [5*] (UMIL) - Preliminary analysis on resistant accessions showed the absence of molecular markers Ren1 and Run1 linked to resistance traits for American and Asian Vitisspecies. Resistant genotypes will be assessed in vitro for tolerance to salinity and drought [6*](ARC) In olive: - more than #450 accessions from different collections, including Tunisian, Algerian and Moroccan germplasms will be tested for tolerance/susceptibility to high-temperature stress on flowers, soil salinity and drought stress, as well as for verticillium wilt and olive fruit fly (IOSV, INRA-MO, ENSA and IO) [8*] FREECLIMB PRIMA Meknes 2019

  8. WP1 - Phenotyping for traits relevant to resilient fruit systems production T1.2 Phenotypingsegregating progenies to deciphergeneticdeterminismfortraitsrelevant to predefinedideotypes. M 1-24. Task Leader: CU. Partnersinvolved: UMIL, CREA, CRAG, INRA-GAFL, INRA-BFP, CU, IO. This task will involve the phenotypic characterization of diverse cross-populations for each species addressed, in order to decipher genetic determinism (WP2), focusing on existing progenies (F1, F2, back-crosses). In citrus: - diploid lemon progenies from C. latipes× C. limonand triploid lemon progenies (C. limonhybrids) will be phenotyped for scion tolerance/resistance to Mal secco[*3] (CREA) - different progenies from Citrus rootstocks, including seedlings from C. latipes× P. trifoliataand seedlings from C. reticulata× P. trifoliata(SxR) [*4] will be phenotyped for drought, salinity stress tolerance and lime-induced iron chlorosis (CU). In grape phenotypic responses to drought and salinity stresses will be assessed in a progeny of 150 individuals from M1 x V. berlandieri(UMIL)[*7]. M1 is a rootstock with reduced vigor and high degree of tolerance to iron chlorosis and salinity. In olive, a segregating progeny from cross Coratina x Chemlaliwill be assessed for olive fruit fly tolerance (IO) [9*]. FREECLIMB PRIMA Meknes 2019

  9. WP1 - Phenotyping for traits relevant to resilient fruit systems production T1.2 Phenotypingsegregating progenies to deciphergeneticdeterminismfortraitsrelevant to predefinedideotypes. M 1-24. Task Leader: CU. Partnersinvolved: UMIL, CREA, CRAG, INRA-GAFL, INRA-BFP, CU, IO. In apricot, seedlings from different progenies having different genetic background will be assessed for resistance/tolerance to blossom brown rot (UMIL) [11*]. In peach x almond: -progenies including BC1 and BC2 almond introgressions in peach will be phenotyped for drought (as a rootstock) and chilling requirements [13*] (CRAG). - a peach x almond training population (#70 individuals, developed by INRA for resistance to pests and pathogens) will be phenotyped for bud phenology, floral induction and flowering quality (floral anomalies) as well as for date of leaves falling (INRA-GAFL). In addition, peach x davidianahybrids progenies will be also phenotyped for the same traits (CRAG) [14*]. In peach (CU), #60 seedlings from progeny Stark Red Gold x Ustun(SRGxU); #40 seedlings from progeny Venus x Ustun (VxU) will be phenotyped for tolerance to drought and calcareous soil. FREECLIMB PRIMA Meknes 2019

  10. WP1 - Phenotyping for traits relevant to resilient fruit systems production T1.3 Environment and managementeffectonadaptivetraits. M 1-24. Task Leader: IPB&GR. Partnersinvolved: UMIL, CRAG, IOSV, IO, INRA-MO, ENSA). This task will apply experimental designs for phenotypic evaluation across countries/environments/management conditions, based on reference collections (peach and olive as model species) enclosing a wide diversity of accessions. AMMI analysis will be used to analyse stability of genotypes across environments. In peach, a RefPop [14*] was designed to optimize comparisons between sites (UMIL, CRAG and IPB&GR). Taking advantage of this design (two replicated blocks), field trials will be conducted under different irrigation regimes (i.e. 100% soil capacity vs rainfed), without disease control management. This will allow to evaluate scion response to irrigation and also the effect of water availability on disease development (e.g. RefPop includes diverse accessions with a range of tolerance/susceptibility to biotic stresses). The recording of climate data and the characterization of the environment in each site will complement phenotypic data allowing detailed studies of GxExM interactions (Task 2.4). In olive, a set of cultivars [8*] common to the four partners (IOSV, INRA-MO, IO and ENSA) will be assessed for abiotic stress resilience (early flowering, heat stress tolerance at bloom, drought) and a set of cultivars common to at least two partners will be assessed for olive fruit fly tolerance. This design over 3 sites will allow studying GxExM interactions and assess the stability of olive cultivars performance across MB. FREECLIMB PRIMA Meknes 2019

  11. WP1 - Phenotyping for traits relevant to resilient fruit systems production T1.4 Omicsanalysis of environmentaladaptationmechanisms. M 12-36. Task Leader: CRAG; Partnersinvolved: UMIL, INRA-MO, ENSA, IPB&GR. Taking advantage of the experimental designs developed under Task 1.3, omics approaches will be applied in peach and olive as model species to analyze how plants adapt to environmental constraints and why certain varieties are tolerant to abiotic and biotic single or multiple stresses. Transcriptomics, proteomics and metabolomics analyses will be used to assess plant responses to different abiotic stress conditions: - drought stress in peach trees - temperature effects on pollen viability in olive A set of genotypes with contrasting behaviour will be chosen and compared based on phenotypic assessments. FREECLIMB PRIMA Meknes 2019

  12. WP1 - Phenotyping for traits relevant to resilient fruit systems production Milestones M1.1- Preliminary list of accessions combining resistance/tolerance to biotic and abiotic stresses M12 Lists shared among partners with supporting comments and data M1.2- Preliminary list of parents to be used in crossing or pre-breeding or to be tested in orchards M18 Lists shared among partners with supporting comments and data M1.3- High-quality transcriptomic, proteomic and metabolomic profiling data for model fruit species M12 Plant tissue samples collected and profiled FREECLIMB PRIMA Meknes 2019

  13. WP1 - Phenotyping for traits relevant to resilient fruit systems production Deliverables D 1.1: Validation of existing protocols for abiotic stress assessment (M24) D 1.2: List of accessions combining resistance/tolerance to biotic and abiotic stresses (M24; update M36) D 1.3: Phenotypic data for genetic analyses provided (M24) D 1.4: New parents of interest to be used in crossing, pre-breeding or to be tested in orchards (M24; update M36) D 1.5: Characterization of adaptive mechanisms signatures in model fruit species (M36) FREECLIMB PRIMA Meknes 2019

  14. WP1 - Phenotyping for traits relevant to resilient fruit systems production Description of work This WP relieson: 1) Extensive germplasm collections already available from project partners: plant material representing spontaneous and domesticated biodiversity will be phenotyped, including collections of wild relatives, local ecotypes and varieties, crossing populations and advanced selections from breeding programs. These materials are expected to be a source of variability for traits relevant to more sustainable and resilient fruit production. Old and new accessions are also included to identify resilient cultivars adapted to climatic scenarios predicted for MB which could i) be proposed as best choices to be promptly planted, ii) serve as donors of resilience/resistance/tolerance traits for the introgression into novel pre-breeding materials. 2) Reliable and already validated methods and protocols for phenotyping the targeted traits that will be spread among the partners to i) promote sharing of knowledge and expertise within the MB and ii) ensure standardized scoring and comparable data. Depending on the trait, phenotyping will be performed in orchard, green/screen-houses, growth chambers or lab, with natural or controlled stress conditions. 3) Already available first-screening phenotypic data for some materials, that will be integrated with additional 2 years records after the start of the project in order to ensure robustness of the results. FREECLIMB PRIMA Meknes 2019

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