Diversity of Phytophthora Species Associated with Quercus ilex L. in Three Spanish Regions Evaluated by NGS

[EN] The diversity of Phytophthora species in declining Fagaceae forests in Europe is increasing in the last years. The genus Quercus is one of the most extended Fagaceae genera in Europe, and Q. ilex is the dominant tree in Spain. The introduction of soil-borne pathogens, such as Phytophthora in Fagaceae forests modifies the microbial community present in the rhizosphere, and has relevant environmental and economic consequences. A better understanding of the diversity of Phytophthora spp. associated with Q. ilex is proposed in this study by using Next Generation Sequencing (NGS) in six Q. ilex stands located in three regions in Spain. Thirty-seven Phytophthora phylotypes belonging to clades 1 to 12, except for clades 4, 5 and 11, are detected in this study, which represents a high diversity of Phytophthora species in holm oak Spanish forests. Phytophthora chlamydospora, P. citrophthora, P. gonapodyides, P. lacustris, P. meadii, P. plurivora, P. pseudocryptogea, P. psychrophila and P. quercina were present in the three regions. Seven phylotypes could not be associated with known Phytophthora species, so they were putatively named as Phytophthora sp. Most of the detected phylotypes corresponded to terrestrial Phytophthora species but aquatic species from clades 6 and 9 were also present in all regions.We would like to thank M. Leon from the Instituto Agroforestal Mediterraneo-UPV (Spain) for its technical assistance. This research was supported by funding from the project AGL2011-30438-C02-01 (Ministerio de Economia y Competitividad, Spain) and Euphresco [Instituto Nacional de Investigacion y Tecnologia Agraria y Agroalimentaria (EUPHESCO-CEP: "Current and Emerging Phytophthoras: Research Supporting Risk Assesssment and Risk Management")].Mora-Sala, B.; Gramaje Pérez, D.; Abad Campos, P.; Berbegal Martinez, M. (2019). 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First report of Cylindrocladiella parva and C. peruviana associated with black-foot disease of grapevine in Spain

Agusti Brisach, C.; Alaniz, S.; Gramaje Pérez, D.; Pérez Sierra, AM.; Armengol Fortí, J.; Landeras, E.; Izquierdo, PM. (2012). First report of Cylindrocladiella parva and C. peruviana associated with black-foot disease of grapevine in Spain. Plant Disease. 96(9):1381-1382. doi:10.1094/PDIS-04-12-0410-PDNS1381138296

High-throughput amplicon sequencing-based analysis of active fungal communities inhabiting grapevine after hot-water treatments reveals unexpectedly high fungal diversity

[EN] The ecology of total fungal communities in grapevine is so far largely derived from studies on culture-dependent methods or cultivation-independent rDNA approaches. Sequencing the ribosomal RNA transcripts (rRNA) would rather reveal the functionally and metabolically active important taxa of the fungal community and provide insights into its activity in the wood. The present study investigated changes in the potentially active fungal communities of internal grapevine wood after Hot-Water Treatment (HWT) in planting material from Czech Republic and Spain at two different times during the propagation process and from two plant zones. We examined fungal communities using both traditional isolation into culture and high-throughput amplicon sequencing (HTAS) of the internal transcribed spacer 2 (ITS2) region in extracted total RNA. HTAS from metatranscriptomic RNA increased the resolution of the fungal community analysis and revealed a highly diverse mycobiota of grapevine wood compared to the traditional method. Fungal diversity differed between grapevine genotypes and showed a temporal variation over the vegetative period. Grapevine planting materials exhibited high fungal diversity after HWT, which demonstrates that the HWT process does not sterilize the internal wood of grapevine. HWT reduced the infection caused by fungal trunk disease pathogens but was not completely effective in eliminating their growth. This study provides important and practically useful insights into the dynamics of active fungal communities in hot-water treated plants, and represents the first study of active fungal communities on grapevine grafted plants by comparing traditional and next-generation sequencing methods. (C) 2018 Elsevier Ltd and British Mycological Society. All rights reserved.This work was supported by Projects No. LD14051 from the Ministry of Education, Youth and Sports, Czech Republic covered by COST Action FA1303, and No. CZ.02.1.01/0.0/0.0/16_025/0007314 "Multidisciplinary research to increase application potential of nanomaterials in agricultural practice" from EFRR (Grant provider: Ministerstvo Skolstvi, mladeze a telovychovy Ceske republiky (MSMT). Access to computing and storage facilities owned by parties and projects contributing to the National Grid Infrastructure MetaCentrum provided under the programme "Projects of Large Research, Development, and Innovations Infrastructures" (CESNET LM2015042), is greatly appreciated. David Gramaje was supported by the DOC-INIA program from the National Institute for Agronomic Research (INIA), co-funded by the European Social Fund. The authors are grateful for the technical assistance of Jesus Alfonso Crespo and Aitor Gavara Vidal.Eichmeier, A.; Pecenka, J.; Penazova, E.; Baranek, M.; Català, S.; León Santana, M.; Armengol Fortí, J.... (2018). High-throughput amplicon sequencing-based analysis of active fungal communities inhabiting grapevine after hot-water treatments reveals unexpectedly high fungal diversity. Fungal Ecology. 36:26-38. https://doi.org/10.1016/j.funeco.2018.07.011S26383

Droplet Digital PCR technology for detection of Ilyonectria liriodendri from grapevine environmental samples

http://apsjournals.apsnet.org/feedback/show[EN] Black-foot disease is one of the most important soilborne diseases affecting planting material in grapevine nurseries and young vineyards. Accurate, early, and specific detection and quantification of black-foot disease causing fungi are essential to alert growers and nurseries to the presence of the pathogens in soil, and to prevent the spread of these pathogens through grapevines using certified pathogen-free planting material and development of resistance. We comparatively assessed the accuracy, efficiency, and specificity of droplet digital PCR (ddPCR) and real-time PCR (qPCR) techniques for the detection and quantification of Ilyonectria liriodendri in bulk and rhizosphere soils, as well as grapevine endorhizosphere. Fungal abundance was not affected by soil-plant fractions. Both techniques showed a high degree of correlation across the samples assessed (R2 = 0.95) with ddPCR being more sensitive to lower target concentrations. Roots of asymptomatic vines were found to be a microbial niche that is inhabited by black-foot disease fungi.David Gramaje was supported by the DOC-INIA program from the INIA, co-funded by the European Social Fund. María del Pilar Martínez-Diz was supported by the FPI-INIA program from the INIA.Martínez Martínez, MDP.; Andrés-Sodupe, M.; Berbegal Martinez, M.; Bujanda, R.; Díaz-Losada, E.; Gramaje Pérez, D. (2020). Droplet Digital PCR technology for detection of Ilyonectria liriodendri from grapevine environmental samples. Plant Disease. 104(4):1144-1150. https://doi.org/10.1094/PDIS-03-19-0529-RES11441150104

New Phaeoacremonium species isolated from sandalwood trees in Western Australia

Thirty-eight Phaeoacremonium isolates collected from pruning wounds of tropical sandalwood in Western Australia were studied with morphological and cultural characteristics as well as phylogenetic analyses of combined DNA sequences of the actin and beta-tubulin genes. Three known Phaeoacremonium species were found, namely P. alvesii, P. parasiticum, and P. venezuelense. Phaeoacremonium venezuelense represents a new record for Australia. Two new species are described: P. luteum sp. nov. can be identified by the ability to produce yellow pigment on MEA, PDA, and OA, the predominance of subcylindrical to subulate type II phialides, and the mycelium showing prominent exudate droplets observed as warts; and P. santali sp. nov. which can be separated from other species producing pink colonies on MEA by the predominance of type I and II phialides, the distinct brownish olive colonies in OA, and slow growth.We acknowledge Pablo Castillo (IAS-CSIC) for sharing the equipment to perform microscopic observations. Isolations were made under project PRJ-004677 "Heartwood Rot Identification and Impact in Sandalwood (Santalum album)" funded by the Rural Industries Research and Development Corporation of Australia. We thank Len Norris and Diane White for assistance with collection and isolation.Gramaje Pérez, D.; León Santana, M.; Pérez Sierra, AM.; Burgess, T.; Armengol Fortí, J. (2014). 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Fungal trunk pathogens associated with wood decay of almond trees on Mallorca (Spain)

Severe decline of almond trees has recently been observed in several orchards on the island of Mallorca (Balearic Islands, western Mediterranean Sea). However, the identity of the causal agents has not yet been investigated. Between August 2008 and June 2010, wood samples from branches of almond trees showing internal necroses and brown to black vascular streaking were collected in the Llevant region on the island of Mallorca. Several fungal species were subsequently isolated from the margin between healthy and symptomatic tissue. Five species of Botryosphaeriaceae (namely Botryosphaeria dothidea, Diplodia olivarum, D. seriata, Neofusicoccum australe and N. parvum), Eutypa lata, Phaeoacremonium iranianum and Phomopsis amygdali were identified based on morphology, culture characteristics and DNA sequence comparisons. Neofusicoccum parvum was the dominant species, followed by E. lata, D. olivarum and N. australe. First reports from almond include D. olivarum and Pm. iranianum. Two species are newly described, namely Collophora hispanica sp. nov. and Phaeoacremonium amygdalinum sp. nov.We thank 'Conselleria d'Agricultura, Mediambient i Territori' of the Balearic Islands for funding this research. We appreciate the helpful assistance in field works from technicians and land owners: Tomeu Melis, Toni Nicolau, Beatriz Blanquer and Alejandro Aristondo. The authors also wish to thank the staff of 'Conselleria d'Agricultura, Mediambient i Territori': Alicia Nieto, Xavier Nadal and Toni Martorell. We would like to thank Walter Gams for kindly assisting with the nomenclature for the new species.Gramaje Pérez, D.; Agusti Brisach, C.; Pérez Sierra, AM.; Moralejo, E.; Olmo, D.; Mostert, L.; Damm, U.... (2012). Fungal trunk pathogens associated with wood decay of almond trees on Mallorca (Spain). Persoonia: a mycological journal. 28:1-13. doi:10.3767/003158512X626155S1132

Challenges of viticulture adaptation to global change: tackling the issue from the roots

[EN] Viticulture is facing emerging challenges not only because of the effect of climate change on yield and composition of grapes, but also of a social demand for environmental-friendly agricultural management. Adaptation to these challenges is essential to guarantee the sustainability of viticulture. The aim of this review is to present adaptation possibilities from the soil-hidden, and often disregarded, part of the grapevine, the roots. The complexity of soil-root interactions makes necessary a comprehensive approach taking into account physiology, pathology and genetics, in order to outline strategies to improve viticulture adaptation to current and future threats. Rootstocks are the link between soil and scion in grafted crops, and they have played an essential role in viticulture since the introduction of phylloxera into Europe at the end of the 19th century. This review outlines current and future challenges that are threatening the sustainability of the wine sector and the relevant role that rootstocks can play to face these threats. We describe how rootstocks along with soil management can be exploited as an essential tool to deal with the effects of climate change and of emerging soil-borne pests and pathogens. Moreover, we discuss the possibilities and limitations of diverse genetic strategies for rootstock breeding.This work is framed in the networking activities of RedVitis (AGL2015-70931-REDT) and RedVitis 2.0 (AGL2017-90759-REDT), funded by the State Research Agency (AEI) of the Spanish Ministry of Science and Innovation. Ms Diana Marin is beneficiary of postgraduate scholarship funded by Universidad Publica de Navarra (FPI-UPNA-2016). Dr Juan Emilio Palomares-Rius acknowledges the State Research Agency (AEI) of the Spanish Ministry of Science and Innovation for the 'Ramon y Cajal' Fellowship RYC-2017-22228 and Dr David Gramaje acknowledges Spanish Ministry of Economy and Competitiveness for the 'Ramon y Cajal' Fellowship RYC-2017-23098.Marín, D.; Armengol Fortí, J.; Carbonell-Bejerano, P.; Escalona, J.; Gramaje Pérez, D.; Hernández-Montes, E.; Intrigliolo, DS.... (2021). Challenges of viticulture adaptation to global change: tackling the issue from the roots. Australian Journal of Grape and Wine Research. 27(1):8-25. https://doi.org/10.1111/ajgw.12463S825271AGÜERO, C. B., URATSU, S. L., GREVE, C., POWELL, A. L. T., LABAVITCH, J. M., MEREDITH, C. P., & DANDEKAR, A. M. (2005). Evaluation of tolerance to Pierce’s disease andBotrytisin transgenic plants ofVitis viniferaL. expressing the pear PGIP gene. Molecular Plant Pathology, 6(1), 43-51. doi:10.1111/j.1364-3703.2004.00262.xAgustí-Brisach, C., Mostert, L., & Armengol, J. (2013). Detection and quantification ofIlyonectriaspp. associated with black-foot disease of grapevine in nursery soils using multiplex nested PCR and quantitative PCR. Plant Pathology, 63(2), 316-322. doi:10.1111/ppa.12093Agustí-Brisach, C., Gramaje, D., García-Jiménez, J., & Armengol, J. (2013). Detection of black-foot disease pathogens in the grapevine nursery propagation process in Spain. European Journal of Plant Pathology, 137(1), 103-112. doi:10.1007/s10658-013-0221-8Alaniz, S., García-Jiménez, J., Abad-Campos, P., & Armengol, J. (2010). Susceptibility of grapevine rootstocks to Cylindrocarpon liriodendri and C. macrodidymum. Scientia Horticulturae, 125(3), 305-308. doi:10.1016/j.scienta.2010.04.009Alaniz, S., Armengol, J., León, M., García-Jiménez, J., & Abad-Campos, P. (2009). Analysis of genetic and virulence diversity of Cylindrocarpon liriodendri and C. macrodidymum associated with black foot disease of grapevine. Mycological Research, 113(1), 16-23. doi:10.1016/j.mycres.2008.07.002Albacete, A., Martinez-Andujar, C., Martinez-Perez, A., Thompson, A. J., Dodd, I. C., & Perez-Alfocea, F. (2015). Unravelling rootstockxscion interactions to improve food security. Journal of Experimental Botany, 66(8), 2211-2226. doi:10.1093/jxb/erv027Aragüés, R., Medina, E. T., Zribi, W., Clavería, I., Álvaro-Fuentes, J., & Faci, J. (2014). Soil salinization as a threat to the sustainability of deficit irrigation under present and expected climate change scenarios. Irrigation Science, 33(1), 67-79. doi:10.1007/s00271-014-0449-xBarrios-Masias, F. H., Knipfer, T., Walker, M. A., & McElrone, A. J. (2019). Differences in hydraulic traits of grapevine rootstocks are not conferred to a common Vitis vinifera scion. 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Diaporthe diversity and pathogenicity revealed from a broad survey of grapevine diseases in Europe

[EN] Species of Diaporthe are considered important plant pathogens, saprobes, and endophytes on a wide range of plant hosts. Several species are well-known on grapevines, either as agents of pre-or post-harvest infections, including Phomopsis cane and leaf spot, cane bleaching, swelling arm and trunk cankers. In this study we explore the occurrence, diversity and pathogenicity of Diaporthe spp. associated with Vitis vinifera in major grape production areas of Europe and Israel, focusing on nurseries and vineyards. Surveys were conducted in Croatia, Czech Republic, France, Hungary, Israel, Italy, Spain and the UK. A total of 175 Diaporthe strains were isolated from asymptomatic and symptomatic shoots, branches and trunks. A multi-locus phylogeny was established based on five genomic loci (ITS, tef1, cal, his3 and tub2), and the morphological characters of the isolates were determined. Preliminary pathogenicity tests were performed on green grapevine shoots with representative isolates. The most commonly isolated species were D. eres and D. ampelina. Four new Diaporthe species described here as D. bohemiae, D. celeris, D. hispaniae and D. hungariae were found associated with affected vines. Pathogenicity tests revealed D. baccae, D. celeris, D. hispaniae and D. hungariae as pathogens of grapevines. No symptoms were caused by D. bohemiae. This study represents the first report of D. ambigua and D. baccae on grapevines in Europe. The present study improves our understanding of the species associated with several disease symptoms on V. vinifera plants, and provides useful information for effective disease management.This research was supported by the European COST Action FA1303 on Sustainable control of grapevine trunk diseases (ManaGTD). Surveys and fungal collection performed in Hungary were supported by Szechenyi 2020 programme, the European Regional Development Fund, the Hungarian Government (GINOP-2.3.2-15-2016-00061) and EU H2020 project no. 652601.Guarmaccia, V.; Groenewald, JZ.; Woodhall, J.; Armengol Fortí, J.; Cinelli, T.; Eichmeier, A.; Ezra, D.... (2018). Diaporthe diversity and pathogenicity revealed from a broad survey of grapevine diseases in Europe. Persoonia - Molecular Phylogeny and Evolution of Fungi. 40:135-153. https://doi.org/10.3767/persoonia.2018.40.06S1351534

Effects of hot-water treatment, post-hot-water-treatment cooling and cold storage on the viability of dormant grafted grapevines under field conditions

Background and Aims: Hot-water treatment (HWT) is an effective control method for black-foot and Petri disease, in grapevine propagating material. However, plant hydration and cold storage have emerged as critical factors in the production of quality planting material. The effects of HWT protocols on the performance of dormant plants ready to be sold to producers under ¿eld conditions were investigated. Methods and Results: The effects of HWT at 53°C for 30 min, cooling (post-HWT cooling or no post-HWT cooling) and cold storage (0, 1, 2 and 4 weeks) on sprouting, and shoot length and weight in dormant grafted plants (Tempranillo cultivar grafted onto 110 Richter rootstock) were evaluated. Eight bundles of ten cuttings were treated for each factor combination, and eight additional bundles of ten untreated cuttings were prepared as controls (no HWT). Dormant grafted plants were immediately planted in two ¿eld sites in March 2010. The number of plants that emerged from dormancy was counted in July 2010. In January 2011, shoot length and fresh weight were evaluated. Although signi¿cant, the percentages of plants emerging from dormancy among treatments were relatively small. A signi¿cant reduction was observed in shoot length and weight for all treatments compared with the control, particularly in all variables for non-hydrated hot-water-treated cuttings kept in cold storage for 4 weeks immediately after HWT. Conclusions: The ¿ndings obtained in this study indicate that long-term cold storage could be detrimental to planting material, especially when plants have not been previously hydrated following HWT. Signi¿cance of the Study: This study represents the ¿rst approach for evaluation of different HWT protocols under ¿eld conditions. It improves the knowledge of the different steps used in the HWT process and provides valuable information about the most reliable protocol that can be used successfully in a commercial situation.This research was financially supported by the Projects RTA2007-00023-C04-03 and RTA2010-00009-C03-03 (Instituto Nacional de Investigacion y Tecnologia Agraria y Alimentaria, INIA, Spain) and the European Regional Development Fund (ERDF). The authors acknowledge V. Garrigues and R. Gandia for technical assistance.Gramaje Pérez, D.; Armengol Fortí, J. (2012). Effects of hot-water treatment, post-hot-water-treatment cooling and cold storage on the viability of dormant grafted grapevines under field conditions. Australian Journal of Grape and Wine Research. 18:158-163. doi:10.1111/j.1755-0238.2012.00185.xS15816318Aroca, Á., Gramaje, D., Armengol, J., García-Jiménez, J., & Raposo, R. (2009). Evaluation of the grapevine nursery propagation process as a source of Phaeoacremonium spp. and Phaeomoniella chlamydospora and occurrence of trunk disease pathogens in rootstock mother vines in Spain. European Journal of Plant Pathology, 126(2), 165-174. doi:10.1007/s10658-009-9530-3Burr, T. J. (1989). Effect of Hot Water Treatment on Systemic Agrobacterium tumefaciens Biovar 3 in Dormant Grape Cuttings. Plant Disease, 73(3), 242. doi:10.1094/pd-73-0242CAUDWELL, A., LARRUE, J., BOUDON-PADIEU, E., & MCLEAN, G. D. (1997). Flavescence dorée elimination from dormant wood of grapevines by hot-water treatment. Australian Journal of Grape and Wine Research, 3(1), 21-25. doi:10.1111/j.1755-0238.1997.tb00112.xFourie, P. H., & Halleen, F. (2002). Investigation on the occurrence of Phaeomoniella chlamydospora in canes of rootstock mother vines. Australasian Plant Pathology, 31(4), 425. doi:10.1071/ap02049Fourie, P. H., & Halleen, F. (2004). Proactive Control of Petri Disease of Grapevine Through Treatment of Propagation Material. Plant Disease, 88(11), 1241-1245. doi:10.1094/pdis.2004.88.11.1241Goheen, A. C. (1973). Association of a Rickettsialike Organism with Pierce’s Disease of Grapevines and Alfalfa Dwarf and Heat Therapy of the Disease in Grapevines. Phytopathology, 63(3), 341. doi:10.1094/phyto-63-341Gramaje, D., García-Jiménez, J., & Armengol, J. (2008). Sensitivity of Petri disease pathogens to hot-water treatmentsin vitro. Annals of Applied Biology, 153(1), 95-103. doi:10.1111/j.1744-7348.2008.00229.xGramaje, D., Armengol, J., Salazar, D., López-Cortés, I., & García-Jiménez, J. (2009). Effect of hot-water treatments above 50°C on grapevine viability and survival of Petri disease pathogens. Crop Protection, 28(3), 280-285. doi:10.1016/j.cropro.2008.11.002Gramaje, D., Alaniz, S., Abad-Campos, P., García-Jiménez, J., & Armengol, J. (2010). Effect of hot-water treatmentsin vitroon conidial germination and mycelial growth of grapevine trunk pathogens. Annals of Applied Biology, 156(2), 231-241. doi:10.1111/j.1744-7348.2009.00382.xHalleen, F., Fourie, P. H., & Crous, P. W. (2007). Control of black foot disease in grapevine nurseries. Plant Pathology, 56(4), 637-645. doi:10.1111/j.1365-3059.2007.01613.xHaviland, D. R., Bentley, W. J., & Daane, K. M. (2005). Hot-Water Treatments for Control of Planococcus ficus (Homoptera: Pseudococcidae) on Dormant Grape Cuttings. Journal of Economic Entomology, 98(4), 1109-1115. doi:10.1603/0022-0493-98.4.1109Probst, C., Jones, E. E., Ridgway, H. J., & Jaspers, M. V. (2011). Cylindrocarpon black foot in nurseries—two factors that can increase infection. Australasian Plant Pathology, 41(2), 157-163. doi:10.1007/s13313-011-0103-5Stonerod, P., & Strik, B. (1996). Hot-water Dipping Eradicates Phylloxera from Grape Nursery Stock. HortTechnology, 6(4), 381-383. doi:10.21273/horttech.6.4.381Waite , H. 2002 The effects of hot water treatment on grapevine propagating and planting materia

Fungal pathogens associated with grapevine trunk diseases in Iran

[EN] During 2004-2007 various own rooted vineyards were inspected to study the fungi associated with vine trunk diseases in Iran. Samples from declining vines showing yellowing and reduced growth and different symptoms in wood, including browning of the wood, necrosis, brown and black streaking and white rot were collected. Fungal isolations were made from affected tissues onto Malt Extract Agar (MEA) supplemented with 1 g l(-1) streptomycin sulphate (MEAS). Based on morphological and molecular characteristics, the following species were identified: Phaeoacremonium (Pm.) aleophilum, Phaeomoniella (Pa.) chlamydospora and less frequently Pm. parasiticum, Pm. inflatipes, Pm. cinereum, Cylindrocarpon liriodendri, Diplodia seriata and Neofusicoccum parvum. Results of the pathogenicity tests under field conditions showed that Pa. chlamydospora and Phaeoacremonium spp. caused large wood discoloration 10 months after inoculation without any external foliar symptoms. Phaeomoniella chlamydospora caused larger lesions than Phaeoacremonium spp. All inoculated species were re-isolated from the margin of the lesions completing Koch's postulates. This study represents the first comprehensive work that investigates the molecular and morphological identification and pathogenicity of Phaeoacremonium spp. and Pa. chlamydospora associated with vine decline in Iran. This is also the first report of Pm. inflatipes, N. parvum and D. seriata associated with grapevine decline in this country.Mohammadi, H.; Banihashemi, Z.; Gramaje Pérez, D.; Armengol Fortí, J. (2013). Fungal pathogens associated with grapevine trunk diseases in Iran. Journal of Agricultural Science And Technology. 15(1):137-150. http://hdl.handle.net/10251/91876S13715015