1H-NMR Metabolite Fingerprinting Analysis Reveals a Disease Biomarker and a Field Treatment Response in Xylella fastidiosa subsp. pauca-Infected Olive Trees

Abstract

Xylella fastidiosa subsp. pauca is a xylem-limited bacterial phytopathogen currently found associated on many hectares with the “olive quick decline syndrome” in the Apulia region (Southern Italy), and the cultivars Ogliarola salentina and Cellina di Nardò result in being particularly sensitive to the disease. In order to find compounds showing the capability of reducing the population cell density of the pathogen within the leaves, we tested, in some olive orchards naturally-infected by the bacterium, a zinc-copper-citric acid biocomplex, namely Dentamet®, by spraying it to the crown, once per month, during spring and summer. The occurrence of the pathogen in the four olive orchards chosen for the trial was molecularly assessed. A 1H NMR metabolomic approach, in conjunction with a multivariate statistical analysis, was applied to investigate the metabolic pattern of both infected and treated adult olive cultivars, Ogliarola salentina and Cellina di Nardò trees, in two sampling periods, performed during the first year of the trial. For both cultivars and sampling periods, the orthogonal partial least squares discriminant analysis (OPLS-DA) gave good models of separation according to the treatment application. In both cultivars, some metabolites such as quinic acid, the aldehydic form of oleoeuropein, ligstroside and phenolic compounds, were consistently found as discriminative for the untreated olive trees in comparison with the Dentamet®-treated trees. Quinic acid, a precursor of lignin, was confirmed as a disease biomarker for the olive trees infected by X. fastidiosa subsp. pauca. When treated with Dentamet®, the two cultivars showed a distinct response. A consistent increase in malic acid was observed for the Ogliarola salentina trees, whereas in the Cellina di Nardò trees the treatments attenuate the metabolic response to the infection. To note that in Cellina di Nardò trees at the first sampling, an increase in γ-aminobutyric acid (GABA) was observed. This study highlights how the infection incited by X. fastidiosa subsp. pauca strongly modifies the overall metabolism of olive trees, and how a zinc-copper-citric acid biocomplex can induce an early re-programming of the metabolic pathways in the infected trees.

Soil and Leaf Ionome Heterogeneity in Xylella fastidiosa Subsp. Pauca-Infected, Non-Infected and Treated Olive Groves in Apulia, Italy

“…A higher content of Zn and Cu is conceivable for the repeated spray treatments to the canopy with the biocomplex, whereas the low Na content could be in relation with metabolic pathways of the trees being close to the normality. It was recently shown that the biocomplex induces an early re-programming of the infected trees upon the spray treatments characterized by the loss of all disease biomarkers [27]. On the other hand, the leaf ionome comparison between treated and not infected trees, as well as between the olive groves of the "free" and "infected" areas also show clear differences, resulting in Zn being the most discriminative ion between the two situations and could be used as a putative biomarker for tolerance to X. f. subsp.…”

“…Within this context, we observed that the supply of a zinc-copper-citric acid biocomplex to the canopy of X. fastidiosa subsp. pauca-infected olive trees during spring and summer allowed to reduce both the field symptoms (i.e., leaf and twig die-backs) and pathogen cell densities within the leaves [25] as well as a rapid re-programming of the basic metabolism of the infected tree [26,27].…”

Analysis of Plant Metabolites Damaged in Chemical Accidents Using NMR Spectroscopy

“…Previous studies were used to assign peaks and chemical shift regions. 7,[17][18][19][20] We found that there was no significant change between the three spectra, but there was a difference in peak intensity in the unsaturated fatty acid region near 2 ppm and in the polyunsaturated fatty acid region near 2.8 ppm. When oak leaves are exposed to chemicals, the intensity of the peak is reduced in the aldehyde region of 8-12 ppm.…”

“…To this end, measurement techniques were assessed for analysis of the molecular structure of chemicals in solid plant samples by referring to published papers and databases available in domestic and international journals. [7][8][9][10][11][12][13][14][15][16][17][18][19][20] After a pretreatment method was established, analysis of chemical accident-induced damage to plants was carried out using solution-state NMR, high-resolution magic angle spinning NMR (HR-MAS NMR), and solid-state NMR. Solutionstate NMR shows sharp peaks due to fast tumbling, but can only measure substances that are soluble in solvents and it is important to find a suitable solvent.…”

“…It is intended to contribute to the preparation of objects and methodologies for loss estimation. To this end, measurement techniques were assessed for analysis of the molecular structure of chemicals in solid plant samples by referring to published papers and databases available in domestic and international journals 7–20 . After a pretreatment method was established, analysis of chemical accident‐induced damage to plants was carried out using solution‐state NMR, high‐resolution magic angle spinning NMR (HR‐MAS NMR), and solid‐state NMR.…”