Management of grapevine gene-banks and prevention from virus infections

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Meeting of COST Actions FA1003 & FA0807 Phytoplasmas and viruses

Mobilization and Evaluation 8 9 May 2012, SOFIA (BULGARIA) Management

P., Venerito P., Savino V., Martelli G.P. Istituto di Virologia

1 Meeting of COST Actions FA1003 & FA0807 Phytoplasmas and viruses management in Grapevine Collections for Germplasm Conservation, Mobilization and Evaluation 8 9 May 2012, SOFIA (BULGARIA) Management of grapevine gene-banks and prevention from virus infections La Notte P., Venerito P., Savino V., Martelli G.P. Istituto di Virologia Vegetale CNR Sezione di Bari Centro Ricerca e Sperimentazione in Agricoltura Basile Caramia

2 The risk of virus infection and the control measures to keep unmodified the sanitary status of plants in the experimental fields vary according with the type and value of the material to be retained/evaluated as well as the place of collection and the local climatic conditions varietal collections for different purposes (i.e. rare autochthonous germplasm) collection of accessions deriving from clonal selection programmes collection of parentals for breeding activities comparison/ evaluation plot for clones, varieties, new crosses and candidate new varieties primary sources /initial materials of certified clones

4 Depending on their frequency, distribution, harmfulness and epidemiology, among the high number of viruses reported on grapevine only few represent a real risk for the reinfection and the evaluation in the gene banks. They are those associated with the three most dangerous diseases considered by the European legislation on certification of grapevine propagation material

5 Infectious degeneration disease (Nepoviruses) Leafroll disease Rugose wood disease

6 Fanleaf degeneration is transmitted by: infected propagation material nematode vectors Nematode ( soil mediated) transmission

7 Nematode vettore: Xiphinema index Ordine: Dorylaimida Famiglia: Grapevine Longidoridae fanleaf virus and its specific vector Molto diffuso in Italia

8 16 viruses (11 Europe) GFLV Grapevine fanleaf virus ArMV Arabis mosaic virus SLRSV Strawberry latent ringspot virus AILV Artichoke Italian latent virus TBRV Tomato blackring virus RpRSV Raspberry ringspot virus GCMV Grapevine chrome mosaic virus GBLV Grapevine Bulgarian latent virus GTRSV Grapevine Tunisian ringspot virus CLRV Cherry leafroll virus GARSV Grapevine Anatolian ringspot virus GDefV Grapevine deformation virus PRMV Peach rosette mosaic virus TRSV Tobacco ringspot virus ToRSV Tomato ringspot virus BLMoV Blueberry leaf mottle virus EU, California EU D, IT, TK, PT IT D, YU, IL, TK, CA D HU-CZ-CR-AU BG, HU, SR TU D TK TK Xiphinema index, (X. italiae and X. vuittenezi?) Xiphinema diversicaudatum Longidorus apulus (only on vegetables) Longidorus attenuatus Paralongidorus maximus Unknown (X. index?) unknown unknown unknown unknown unknown Xiphinema americanum Xiphinema americanum, Longidorus diadecturus, L. elongatus Xiphinema americanum, X. bricolensis, rivesi, californicum, targanense, intermedium unknown

9 Transmission of leafroll and rugose wood by not flying insects vectors

Mealybugs and soft scales GVA: Planococcus citri, Pl. ficus, Pseudococcus longispinus, Ps. affinis, Ps. comstocki Neopulvinaria innumerabilis GVB: Pl. ficus, Ps. longispinus Ps.

10 Mealybugs and soft scales GVA: Planococcus citri, Pl. ficus, Pseudococcus longispinus, Ps. affinis, Ps. comstocki Neopulvinaria innumerabilis GVB: Pl. ficus, Ps. longispinus Ps. affinis, GLRaV-1: Helicoccus bohemicus, Phenacoccus aceris Parthenolecanium corni, Neopulvinaria innumerabilis GLRaV-3: Planococcus ficus, Pl. citri Pseudococcus longispinus. Ps. affinis, Ps. calceolariae, Ps.. viburni, Ps. maritimus Ps comstocki, Pulvinaria vitis GLRaV-5: Ps. longispinus GLRaV-9. Ps. longispinus

11 Mealybugs are much more efficient than soft scale in vectoring grapevine viruses in open field can transmit and are mobile in all stages have a high number of generation x year due to their behavior are difficult to control the transmission is semipersistent and not very specific. It allows the spread of infection with low population levels as well as the survival of a limited number of adults (hidden under the bark) is able to renew severe infestations.

12 Once planted a new vineyard the grower has no instruments to repair the possible damages caused by viruses The control of virus infection is based on the PREVENTION

13 Selection and preparation of primary sorces Official registration of clones Conservation for the premultiplication Pre-multiplication in vivo or in vitro Mmultiplication CONTROLLI Servizio Controllo Vivai

14 Sanitation Thermoterapy Meristem tip culture Meristem tip culture

15 PREVENTIVE MEASURES TO BE IMPLEMENTED BEFORE THE ESTABLISHMENT OF A NEW EXPERIMENTAL VINEYARD Use of propagation material (rootstock and scion) healty or controlled to avoid the introduction of new viruses for the area Use of soil free from nematodes vectors (and fungal root rot agents) Control of soil-borne vectors

16 Controls to carry out before planting Presence of rootstock shoots (also along the boundaries) of any preceding vineyard Nematological analysis (only vector species) mycological analysis (i.e. Armillaria mellea, Rosellinia necatrix)

17 Chemical control of soil-borne vectors? Fumigation Nematocides LOW EFFICACY IN DEPTH, VERY HIGH COSTS, HIGH ENVIRONMENTAL IMPACT

Fig Pistachio ELIMINATION OF ALTERNATIVE HOST SPECIES Ficus carica (Fig) Pistacia vera (Pistachio) Ceratonia siliqua (Carob tree) Prunus domestica (European Plum) Citrus sp.

(Pepper) Fragaria X ananassa (Strawberry) Solanum lycopersicum (Tomato) Solanum tuberosum (Potato) Petunia X hybrida (Petunia) Chenopodium amaranticolor Solanum nigrum X.

18 Fig Pistachio ELIMINATION OF ALTERNATIVE HOST SPECIES Ficus carica (Fig) Pistacia vera (Pistachio) Ceratonia siliqua (Carob tree) Prunus domestica (European Plum) Citrus sp. (Citrus) Malus pumila (Apple) Olea europaea (Olive) Salix sp. (Willow) Brassica oleracea Mill. (Broccoli) Capsicum frutescens L. (Pepper) Fragaria X ananassa (Strawberry) Solanum lycopersicum (Tomato) Solanum tuberosum (Potato) Petunia X hybrida (Petunia) Chenopodium amaranticolor Solanum nigrum X. diversicaudatum: Apricot, Avocado, Black Currant, Blackberry, Carnation, Celery, Citrus, Juniper, Cucumber, Fig, Garden Raspberry, Nectarine, Okra, Peanut, Pear, Perennial Ryegrass, Petunia, Rose, Soybean, Strawberry, Tomato

Correct grubbing up and replanting of vineyard Use

Uproot the vines carefully removing and burning all

cm deep plowing Land left for at least 3 years using

19 Correct grubbing up and replanting of vineyard Use of systemic herbicides to kill the vines (optional) Uproot the vines carefully removing and burning all the roots (with excavator and hand finishing) cm deep plowing Land left for at least 3 years using vernine cover crops (cereals or fodder) Summer plowings

vineyards infected with infectious degeneration and infested by nematode vectors

20 - Correct choice of field location (possibly isolated and not placed under the slope of other vineyards) - Leave a band edge at least of 10 m from other bordering vineyards infected with infectious degeneration and infested by nematode vectors -Ensure the isolation of surface water from sourrounding vineyards (i.e. digging a ditch)

21 Isolation of premultiplication fields and mother plant blocks

Use dedicated equipments or clean them carefully from soil from other fields - Avoid to

22 AND AFTER PLANTING.. Measures, instruments and facilities to prevent or control the reinfection /over-infection - Use dedicated equipments or clean them carefully from soil from other fields - Avoid to introduce in the gene-banks material infected by nepoviruses transmissible by nematodes - Monitoring continuously the possible occurrence of new symptoms - Avoid to overgraft the plants

23 The populations of mealybugs must be continuously and carefully monitored especially when favorable climatic conditions occur Finding very low infestation levels is difficult and therefore the infestation may go unnoticed for several years peeling away bark on trunks checking where the adults use to over-winter checking on the roots at the base of vine trunk using specific pheromone traps to attract (in surrounding vineyards up to 400 m) and capture the males (probably now the best system to determine low infestation rates)

24 Insecticide for chemical control dormant organophosphate insecticide (i.e. chlorpirifos) in-season application of oragophosphate or carbammate (i.e. methomyl) insect growth regulators i.e. buprofezin (Applaud) nicotinoid systemic insecticide (imidacloprid) N of application, time of treatment, distribution by drip irrigation (in open field) or foliar treatment (mainly on plant in pots)

26 Facilities

27 the surveillance should be very high especially for the mealybugs but their control is cheaper and much more effective

28 Seed transmission Can the seedling be infected after a cross when the parentals are infected? None of the viruses associate with leafroll, rugose wood and fleck is reported to be seed-borne GFLV occurs in the pollen and the endosperm but there are conflicting reports on seed transmission. Seed transmission has been reported for most, but not all, nepoviruses, and BLMoV and CLRV are efficiently transmitted by pollen

29 For several viruses the reproduction could be considered as an alternative procedure of sanitation Usually the reinfection of new germplasm occurs in the evaluation plot for the common technique to over-graft adult plants in order to accelerate the fruit set for the first selection

30 Evaluation plot for new candidate varieties It s strongly suggested to keep the original seedlings self-rooted in pots up to the end of the first evaluation results

31 Thanks for the attention

California Seed & Plant Lab., Inc Pleasant Grove Rd, Elverta CA Phone: (916) Fax: (916)

California Seed & Plant Lab., Inc Pleasant Grove Rd, Elverta CA Phone: (916) Fax: (916) Viruses testing RT-PCR for Virus diagnosis We have developed a PCR procedure to detect various strains of each virus. A multiprimer cocktail is used to detect all strains of the virus. For many viruses,