Detection and identification of Phytophthora sp. associated with ink disease of chestnut (Castanea sativa) from natural soils

• Autor(a) principal: Machado, Júlia Melo
• Data de Publicação: 2022
• Tipo de documento: Dissertação
• Idioma: por
• Título da fonte: Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
• Texto Completo: http://hdl.handle.net/10198/26454
• topic: Ink disease; Chestnut tree; Phytophthora cinnamomi; Molecular biology; Domínio/Área Científica::Engenharia e Tecnologia::Outras Engenharias e Tecnologias

One of the biggest threats for the chestnut tree in Portugal and in Europe is the ink disease. The disease is caused by two Phytophthora species (Phytophthora cinnamomi and Phytophthora cambivora). These root parasites have biological and epidemiological characteristics that provide them with rapid adaptation to the environmental conditions and have a high capacity for survival. With this work we intend to develop molecular methods for the detection and identification of Phytophthora in chestnut soils, providing greater sensitivity of detection and greater specificity of identification. In this study, two methods were compared for the detection of Phytophthora from soils infected with chestnut ink disease (natural soil from a stand with diseased chestnut trees and potting soil from greenhouses). In the classical method (baiting method), for the detection of Phytophthora, leaf discs of chestnut (Castanea sativa) were used and subsequent identification of isolates. For molecular methods, soil DNA was extracted from the DNeasy® PowerSoil® Pro Kit (Qiagen) and was amplified from the ITS region of ribosomal DNA (rDNA) using the universal primers ITS4 and ITS6. It was possible to obtain 67% of positive detection from natural soils by baiting method and 75% of positive detection from soil of pots from greenhouse. P. cinnamomi was the species identified from the baiting method and the nucleotide identity of the ITS region of the three identified isolates ranged between 98 and 100%. Trap tissue assays are very useful when the aim is to obtain isolates of different Phytophthora species and DNA-PCR assays are valuable and basic requirements for integrated management programs to evaluate the control measures applied to minimize impacts and control the disease.