Table 1.
Bacterial strains, mutant and plasmids used in this study.
Table 2.
Quali-quantitative HPLC/DAD/MS analysis of FO, FC, VN and TV extracts.
Table 3.
Effects on bacterial growth, and on the trans-activation of hrpA and psnI promoters of the polyphenolic extracts and their main constituents.
Fig 1.
Pathogenicity test with Psn23 on oleander explants, following treatment with polyphenolic extracts VN, TV, FO or FC.
Explants from adult oleander plants were inoculated with P. savastanoi pv. nerii strain Psn23, in the presence or absence of the VN, TV, FO or FC extracts (100 μM). As negative control the non pathogenic mutant ∆hrpA was used. (A) Development of hyperplastic knots at 21 dpi with (from left to right): Psn23, ∆hrpA, Psn23+VN, Psn23+TV, Psn23+FO, Psn23+FC. The symptoms are detectable as swelling at the inoculated end of oleander explants. (B) Normalized weight increase of oleander explants at 21 dpi inoculated with (from left to right): Psn23, ∆hrpA, Psn23+VN, Psn23+TV, Psn23+FO, Psn23+FC. Values are means ± SD of nine replicates for each treatment. Different letters indicate significant differences among means at P < 0.05, according to Tukey's test.
Fig 2.
Relative gene expression analysis of key genes correlated to TTSS and QS of Psn23.
Relative mRNA levels of hrpA, hrpL, hrpV, hrpRS, rpoN, lon, psnI, psnR genes of Psn23, grown in MM supplemented with the polyphenolic extracts TV, VN, FO or FC compared to levels in MM alone (untreated). The data are expressed as the average of three replicates ± SD. Asterisks indicate significant differences compared with the untreated sample at P <0.05.
Fig 3.
Effect of polyphenolic extracts on Congo red dye absorption of Psn23 bacterial cultures.
Percentage of Congo red dye absorption of Psn23 bacterial cultures, grown in MM amended with the polyphenolic extracts VN, TV, FO or FC. The data were calculated according to the formula: [(Xunk−XΔhrpA) / (XWT−XΔhrpA)] *100 where XWT and XΔhrpA are the ratio OD490/OD600 for Psn23 and ΔhrpA respectively. The data represent the means ± SD of three replicates. All treatments are statistically significant (P <0.05).
Fig 4.
ELISA assay on Psn23 bacterial supernatant amended with polyphenolic extracts.
Quantification of HrpA protein by ELISA assay on bacterial supernatant of wild type Psn23 grown on MM, or on MM amended with the polyphenolic extracts VN, TV, FO, or FC. As a negative control the ∆hrpA mutant was used. As a reference for quantification, a standard curve was established by a serial dilution of the Psn23 HrpA recombinant protein (117 pg/ml– 40 ng/ml). The data represent the means ± SD of three replicates. Statistically significant differences are represented by different letters above the bars (ANOVA and Tukey’s test, P < 0.05).
Fig 5.
Current measurements on SR vesicles adsorbed on a SSM.
Current signals induced by 100 μM ATP concentration jumps in the presence of 10 μM Ca2+free and in the absence (black curve, control measurement) or in the presence of CuCl2 (red curve) or of the polyphenolic compounds EGCG, catechin, oleuropein, hydroxytyrosol and chlorogenic acid (green curves).