Browsing by Author "Balestra, G. M."
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- Chemical characterization and antimicrobial activity of 6 plant essential oils against Pseudomonas syringae pv. actinidiae and P. syringae pv. actinidifoliorumPublication . Silva, M. Nunes da; Machado, J.; Mazzaglia, A.; Balestra, G. M.; Vasconcelos, M. W.; Carvalho, S. M. P.Introduction: Kiwifruit bacterial canker (KBC), caused by Pseudomonas syringae pv. actinidiae (PSA), is currently the most destructive disease of kiwifruit worldwide. It was introduced in Portugal in 2010, leading to drastic production losses and plant mortality. Conversely, a closely related pathovar, P. syringae pv. actinidifoliorum (PSAF), only causes necrotic spots, not being associated with plant mortality. Current control techniques are expensive, consisting in the application of copper- and antibiotics-based compounds, with serious risks of environmental contamination and development of resistant bacteria. Therefore, the aim of this work was to characterize six plant essential oils against (anise, basil, cardamom, cumin, fennel and laurel) by gas chromatography–mass spectrometry (GC-MS) and evaluate their antimicrobial potential against different strains of PSA and PSAF. Methods: 1. Two highly virulent PSA strains (CFBP 7286 and 1F, isolated from Italy and France, respectively) and two less virulent PSAF strains (ICMP 18804 and ICMP 19441, isolated from New Zealand and Australia, respectively) were grown overnight in LB broth at 27 ᵒC and with shaking. 2. Bacterial inoculum (1-2x107 CFU.mL-1) was plated onto Nutrient Sucrose Agar medium and 20 μL droplets of each essential oil concentration were placed in 5 mm diameter paper discs (N = 3). 3. Plates were incubated at 28 ᵒC for 48 h, after which the inhibition zone was measured. 4. Essential oils chemical characterization was performed by GC-MS. Conclusions: Plant essential oils differed in their composition; however, compounds such as 3-carene, eucalyptol, camphor, fenchone and 4-terpineol were detected transversely in all oils. Although cumin presented the most simple composition, it showed MIC as low as 0.1 % for PSA 19441. In general, basil, cardamom and fennel had higher MICs, whereas anise had the lowest. PSAF strain 19441 was the most sensitive to almost all essential oils tested.
- Early pathogen recognition and antioxidant system activation contributes to actinidia arguta tolerance against pseudomonas syringae pathovars actinidiae and actinidifoliorumPublication . Nunes da Silva, M.; Vasconcelos, M. W.; Gaspar, M.; Balestra, G. M.; Mazzaglia, A.; Carvalho, Susana M. P.Actinidia chinensis and A. arguta have distinct tolerances to Pseudomonas syringae pv. actinidiae (Psa), but the reasons underlying the inter-specific variation remain unclear. This study aimed to integrate the metabolic and molecular responses of these two kiwifruit species against the highly pathogenic Psa and the less pathogenic P. syringae pv. actinidifoliorum (Pfm) bacterial strains. Disease development was monitored weekly till 21 days post inoculation (dpi), analysing a broad number and variety of parameters including: colony forming units (CFU), foliar symptoms, total chlorophylls, lipid peroxidation, soluble polyphenols, lignin and defense-related gene expression. At the end of the experimental period A. chinensis inoculated with Psa presented the highest endophytic bacterial population, whereas A. arguta inoculated with Pfm showed the lowest values, also resulting in a lower extent of leaf symptoms. Metabolic responses to infection were also more pronounced in A. chinensis with decreased total chlorophylls (up to 55%) and increased lipid peroxidation (up to 53%), compared with non-inoculated plants. Moreover, at 14 dpi soluble polyphenols and lignin concentrations were significantly higher (112 and 26%, respectively) in Psa-inoculated plants than in controls, while in A. arguta no significant changes were observed in those metabolic responses, except for lignin concentration which was, in general, significantly higher in Psa-inoculated plants (by at least 22%), comparing with control and Pfm-inoculated plants. Genes encoding antioxidant enzymes (SOD, APX and CAT) were upregulated at an earlier stage in Psa-inoculated A. arguta than in A. chinensis. In contrast, genes related with phenylpropanoids (LOX1) and ethylene (SAM) pathways were downregulated in A. arguta, but upregulated in A. chinensis in the later phases of infection. Expression of Pto3, responsible for pathogen recognition, occurred 2 dpi in A. arguta, but only 14 dpi in A. chinensis. In conclusion, we found that A. arguta is more tolerant to Psa and Pfm infection than A. chinensis and its primary and secondary metabolism is less impacted. A. arguta higher tolerance seems to be related with early pathogen recognition, the activation of plant antioxidant system, and to the suppression of ET and JA pathways from an earlier moment after infection.
- Evaluation of the susceptibility of actinidia chinensis var. deliciosa and a. arguta var. arguta to pseudomonas syringae pv. actinidiaePublication . Silva, M. Nunes da; Vasconcelos, M. W.; Mazzaglia, A.; Balestra, G. M.; Carvalho, S. M. P.
- Exploring the expression of defence-related genes in Actinidia spp. after infection with Pseudomonas syringae pv. actinidiae and pv. actinidifoliorum: first stepsPublication . Silva, M. Nunes da; Machado, J.; Balestra, G. M.; Mazzaglia, A.; Vasconcelos, M. W.; Carvalho, S. M. P.Kiwifruit bacterial canker (KBC), caused by Pseudomonas syringae pv. actinidiae (PSA), is currently the most destructive disease of kiwifruit worldwide. Conversely, a closely related bacterial strain, P. syringae pv. actinidifoliorum (PFM), only causes necrotic spots and has not been associated with plant mortality. Moreover, there is some evidence on the higher susceptibility of the Actinidia chinensis var. deliciosa kiwifruit species to KBC, compared with A. arguta, but the reasons behind it are still largely unknown. In this work, micropropagated plants of Actinidia chinensis var. deliciosa 'Hayward' and A. arguta var. arguta 'Ken's Red' were inoculated with PSA or with PFM (10(7) CFUs mL(-1)). Disease development was monitored 1, 2 and 5 days post inoculation (dpi) through the determination colony forming units (CFUs) and the expression analysis of six plant defence-related genes (APX, CAT, SOD, LOX1, SAM and TLP1). At 5 dpi, CFUs in plant tissues inoculated with PSA and PFM were, respectively, 17.4-fold and 2.8-fold higher in A. chinensis compared with A. arguta. Expression of antioxidant enzyme-related genes was very distinct between the two kiwifruit species: SOD expression was drastically increased in A. chinensis (up to 2.1-fold, 5 dpi), whereas in A. arguta CAT was the most upregulated gene (up to 1.7-fold, 2 dpi). LOX1, involved in jasmonic acid biosynthesis, was upregulated in both species, however reaching the highest values at 2 dpi in A. chinensis (2.2 fold) and 1 dpi in A. arguta (1.9-fold). It is concluded that A. arguta is much more tolerant to PSA than A. chinensis and that the molecular mechanisms between the two kiwifruit species involve specific defence pathways being triggered at distinct moments after plant infection.
- Kiwifruit bacterial canker: novel insights on an old problemPublication . Silva, M. Nunes da; Machado, J.; Balestra, G. M.; Mazzaglia, A.; Vasconcelos, M. V.; Carvalho, S. M. P.Framework: Kiwifruit bacterial canker (KBC), caused by Pseudomonas syringae pv. actinidiae (PSA), is currently the most destructive disease of kiwifruit worldwide. It was introduced in Portugal in 2010, inducing production losses up to 80 %. Conversely, a closely related pathovar, P. syringae pv. actinidifoliorum (PSAF), only causes necrotic spots, not being associated with plant mortality. There is some evidence on the higher susceptibility of green kiwifruit (Actinidia deliciosa) cultivars to KBC, compared with the berry kiwi (A. arguta), but the reasons behind it are still largely unknown. The aim of this work was to evaluate how kiwi plants with reported distinct tolerance to the disease responded to artificial infection in terms of defence-related genes expression. Methods: Micropropagation of A. deliciosa cv. ‘Hayward’ and A. arguta cv. ‘Ken’s Red’ plants. Plant inoculation with a virulent Pseudomonas syringae pv. actinidiae strain (PSA) and with a low virulent P. syringae pv. actinidifoliorum strain (PSAF). Bacterial colony forming units (CFUs) determination in plant tissues 1, 2 and 5 days post inoculation (dpi). Defence-related gene expression analysis by reverse transcription polymerase chain reactions (qRT-PCR) at 1, 2 and 5 dpi. Conclusions: A. deliciosa cv. ‘Hayward’ seems to be much more susceptible to PSA infection than A. arguta cv. ‘Ken’s Red’. SOD expression was drastically increased in ‘Hayward’, whereas C AT was the most upregulated antioxidant-encoding gene in ‘Ken’s Red’. Jasmonic acid and ethylene precursors, LOX and SAM, were upregulated 2 dpi in ‘Hayward’ and already at 1 dpi in ‘Ken’s Red’. LOX1 and SAM transcriptional levels were higher in PSA-inoculated plants, compared with PSAF.