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Murteira Rico dos Santos Campos, Maria Catarina
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- qPCR as a sensitive tool for detecting Fusarium spp. in tomato plantsPublication . Campos, Maria Doroteia; Varanda, Carla; Patanita, Mariana; Ribeiro, Joana A.; Campos, Catarina; Materatski, Patrick; Albuquerque, André; Osa, Nicolás; Félix, M.R.The need to increase food production together with the demand to reduce the application of synthetic chemicals that have consequences on increasing the carbon footprint and negative impacts on the environment and human health, led to the search for alternative methods to protect plants against pathogens. The use of the highly sensitive real-time quantitative PCR (qPCR) arises as an extremely useful tool for studying various agents of infection in plants, such as fungi, viruses, or bacteria leading to a better control of diseases and limiting the use of chemical defence strategies. Given the high incidence of diseases in tomato plants caused by Fusarium spp., their consequent negative economic impacts, and the fact that most phytosanitary treatments are based on the application of synthetic fungicides, the establishment of a molecular-based tool that enables their early and accurate detection is of great interest. Furthermore, it will provide an additional tool for the screening of resistant plants. In the presented study, a TaqMan®-based qPCR method targeting the Fusarium spp.- specific internal transcribed spacer (ITS) region was developed for the simultaneous detection and quantification of a panoply of Fusarium species that affect tomato. As a proof of principle, the new qPCR assay was used to assess Fusarium spp. contamination of tomato field plants and of plants grown under controlled conditions. qPCR combined with the chemistry of TaqMan® MGB probes represents a highly specific and sensitive detection system, even when low amounts of target DNA are present, as in the case of early plant–fungi interactions.
- Identification of candidate genes involved in olive response to anthracnose for a sustainable disease managementPublication . Inácio, Diogo; Félix, M.R.; Campos, Catarina; Patanita, Mariana; Ribeiro, Joana A.; Varanda, Carla; Materatski, Patrick; Albuquerque, André; Osa, Nicolás; Peixe, A.; Campos, Maria DoroteiaAnthracnose is a disease that affect the olive tree caused by fungi of the genus Colletotrichum. These fungi are responsible to premature fruit drop and a consequent decrease in the oil quality, and also cause defoliation of trees compromising the production of the following years. Currently the best control strategy is based on application of synthetic fungicides, with a regulatory pressure in agriculture worldwide to limit its use. Looking for a sustainable disease management, the present study reports the transcriptional changes of olive genes that encode enzymes directly associated with the maintenance of the balance of oxidative oxygen species (ROS), in response to Colletotrichum sp. infection. The cultivar selected for the studies was ‘Galega vulgar’, known to be extremely susceptible to this disease. Plants used in the experiments were from in vitro culture (to warranty their healthy status), transplanted to pots and maintained under controlled conditions. Leaves of the olive plants were inoculated with a spore suspension of Colletotrichum sp., and leaf samples were collected before fungi inoculation (T0) and at 10 days (T1) and 35 days (T2) after inoculation. Confirmation of the presence of the fungi in inoculated plants was performed following a real-time qPCR approach. The selected target genes for expression analysis were Superoxide dismutase (SOD), Endochitinase_EP3-like (CHI2), Glutathione S-transferase L3-like (TransFL3), Glutathione peroxidase 2 (PEROX2), Glutathione S-transferase THETA 1 (THETA), Glutathione S-transferase DHAR2-like (TransfDHAR), Glutathione peroxidase 5 (PEROX5) and Glutathione peroxidase 8 (PEROX8). Following a qPCR approach, a general up-regulation of the target genes was detected, but only CHI2 revealed a significantly up-regulation (p<0.005) between T0 and T2 in response to Colletotrichum sp. infection, being a promising candidate to be later used in functional analysis. We emphasize the importance of this study for the identification of candidate genes to incorporate new sources of resistance of olive trees to anthracnose with the promotion of the development of sustainable management strategies.