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

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.

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Olive Cultivars Susceptible or Tolerant to Xylella fastidiosa Subsp. pauca Exhibit Mid-Term Different Metabolomes upon Natural Infection or a Curative Treatment

Plants ◽

10.3390/plants10040772 ◽

2021 ◽

Vol 10 (4) ◽

pp. 772

Author(s):

Chiara Roberta Girelli ◽

Laura Del Coco ◽

Federica Angilè ◽

Marco Scortichini ◽

Francesco Paolo Fanizzi

Keyword(s):

1H Nmr ◽

Metabolic Response ◽

Xylella Fastidiosa ◽

Natural Infection ◽

Quinic Acid ◽

Term Treatment ◽

Economic Losses ◽

Metabolic Pattern ◽

Olive Cultivars ◽

Olive Quick Decline Syndrome

Xylella fastidiosa subsp. pauca, is a bacterial phytopathogen associated with the “olive quick decline syndrome” (OQDS) causing severe economic losses to olive groves in Salento area (Apulia, Italy). In a previous work, we analyzed by 1H-NMR the metabolic pattern of naturally infected Ogliarola salentina and Cellina di Nardò susceptible cultivars untreated and treated with a zinc-copper citric acid biocomplex and we observed the treatment related variation of the disease biomarker quinic acid. In this study, we focused also on the Leccino cultivar, known to exhibit tolerance to the disease progression. The 1H-NMR-based metabolomic approach was applied with the aim to characterize the overall metabolism of tolerant Leccino in comparison with the susceptible cultivars Ogliarola salentina and Cellina di Nardò under periodic mid-term treatment. In particular, we studied the leaf extract molecular patterns of naturally infected trees untreated and treated with the biocomplex. The metabolic Leccino profiles were analyzed for the first time and compared with those exhibited by the susceptible Cellina di Nardò and Ogliarola salentina cultivars. The study highlighted a specificity in the metabolic response of the tolerant Leccino compared to susceptible cultivars. These differences provide useful information to describe the defensive mechanisms underlying the change of metabolites as a response to the infection, and the occurrence of different levels of disease, season and treatment effects for olive cultivars.

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Ionomic Differences between Susceptible and Resistant Olive Cultivars Infected by Xylella fastidiosa in the Outbreak Area of Salento, Italy

Pathogens ◽

10.3390/pathogens8040272 ◽

2019 ◽

Vol 8 (4) ◽

pp. 272 ◽

Cited By ~ 10

Author(s):

Giusy D’Attoma ◽

Massimiliano Morelli ◽

Pasquale Saldarelli ◽

Maria Saponari ◽

Annalisa Giampetruzzi ◽

...

Keyword(s):

Inductively Coupled Plasma ◽

Xylella Fastidiosa ◽

Optical Emission ◽

Host Genotype ◽

Bacterial Populations ◽

Inductively Coupled ◽

Outbreak Area ◽

Olive Cultivars ◽

Olive Quick Decline Syndrome ◽

Olive Trees

Olive quick decline syndrome (OQDS) is a devastating disease of olive trees in the Salento region, Italy. This disease is caused by the bacterium Xylella fastidiosa, which is widespread in the outbreak area; however, the “Leccino” variety of olives has proven to be resistant with fewer symptoms and lower bacterial populations than the “Ogliarola salentina” variety. We completed an empirical study to determine the mineral and trace element contents (viz; ionome) of leaves from infected trees comparing the two varieties, to develop hypotheses related to the resistance of Leccino trees to X. fastidiosa infection. All samples from both cultivars tested were infected by X. fastidiosa, even if leaves were asymptomatic at the time of collection, due to the high disease pressure in the outbreak area and the long incubation period of this disease. Leaves were binned for the analysis by variety, field location, and infected symptomatic and infected asymptomatic status by visual inspection. The ionome of leaf samples was determined using inductively coupled plasma optical emission spectroscopy (ICP-OES) and compared with each other. These analyses showed that Leccino variety consistently contained higher manganese (Mn) levels compared with Ogliarola salentina, and these levels were higher in both infected asymptomatic and infected symptomatic leaves. Infected asymptomatic and infected symptomatic leaves within a host genotype also showed differences in the ionome, particularly a higher concentration of calcium (Ca) and Mn levels in the Leccino cultivar, and sodium (Na) in both varieties. We hypothesize that the ionome differences in the two varieties contribute to protection against disease caused by X. fastidiosa infection.

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1H NMR-based metabolomic observation of a two-phased toxic mode of action in Eisenia fetida after sub-lethal phenanthrene exposure

Environmental Chemistry ◽

10.1071/en10094 ◽

2011 ◽

Vol 8 (2) ◽

pp. 105 ◽

Cited By ~ 28

Author(s):

Brian P. Lankadurai ◽

David M. Wolfe ◽

André J. Simpson ◽

Myrna J. Simpson

Keyword(s):

1H Nmr ◽

Mode Of Action ◽

Eisenia Fetida ◽

Nmr Spectra ◽

Metabolic Response ◽

Exposure Concentration ◽

Multivariate Statistical ◽

Contaminant Exposure ◽

Tissue Extracts ◽

1H Nuclear Magnetic Resonance

Environmental contextPhenanthrene is a persistent soil contaminant, whose toxic mode of action in earthworms has not been fully examined. We adopt a metabolomics approach, using 1H nuclear magnetic resonance (NMR) spectroscopy, to measure the response of earthworms to sub-lethal phenanthrene exposure. The results indicate that NMR-based metabolomics may be used to monitor responses to sub-lethal levels of contaminants and to delineate their toxic mode of action. Abstract1H NMR-based metabolomics was used to examine the response of the earthworm Eisenia fetida to sub-lethal phenanthrene exposure. E. fetida were exposed via contact tests to six sub-lethal (below the measured LC50 of 1.6 mg cm–2) concentrations of phenanthrene (0.8–0.025 mg cm–2) for 48 h. Multivariate statistical analysis of the 1H NMR spectra of earthworm tissue extracts revealed a two-phased mode of action (MOA). At exposures below 1/16th of the LC50, the MOA was characterised by a linear correlation between the metabolic response and exposure concentration. At exposures ≥1/16th of the LC50, the metabolic response to phenanthrene appeared to plateau, indicating a distinct change in the MOA. Further data analysis suggested that alanine, lysine, arginine, isoleucine, maltose, ATP and betaine may be potential indicators for sub-lethal phenanthrene exposure. Metabolite variation was also found to be proportional to the exposure concentration suggesting that NMR-based earthworm metabolomics is capable of elucidating concentration-dependent relationships in addition to elucidating the MOA of sub-lethal contaminant-exposure.

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1H-NMR Profiling Shows as Specific Constituents Strongly Affect the International EVOO Blends Characteristics: The Case of the Italian Oil

Molecules ◽

10.3390/molecules26082233 ◽

2021 ◽

Vol 26 (8) ◽

pp. 2233

Author(s):

Francesca Calò ◽

Chiara Roberta Girelli ◽

Federica Angilè ◽

Laura Del Coco ◽

Lucia Mazzi ◽

...

Keyword(s):

1H Nmr ◽

Metabolic Profile ◽

1H Nmr Spectroscopy ◽

Virgin Olive Oil ◽

Extra Virgin Olive Oil ◽

Minor Components ◽

Multivariate Statistical ◽

Olive Oils ◽

The World ◽

Plateau Level

Considering the growing number of extra virgin olive oil (EVOO) producers in the world, knowing the influence of olive oils with different geographical origins on the characteristics of the final blend becomes an interesting goal. The present work is focused on commercial organic EVOO blends obtained by mixing multiple oils from different geographical origins. These blends have been studied by 1H-NMR spectroscopy supported by multivariate statistical analysis. Specific characteristics of commercial organic EVOO blends originated by mixing oils from Italy, Tunisia, Portugal, Spain, and Greece were found to be associated with the increasing content of the Italian component. A linear progression of the metabolic profile defined characteristics for the analysed samples—up to a plateau level—was found in relation to the content of the main constituent of the Italian oil, the monocultivar Coratina. The Italian constituent percentage appears to be correlated with the fatty acids (oleic) and the polyphenols (tyrosol, hydroxytyrosol, and derivatives) content as major and minor components respectively. These results, which highlight important economic aspects, also show the utility of 1H-NMR associated with chemometric analysis as a powerful tool in this field. Mixing oils of different national origins, to obtain blends with specific characteristics, could be profitably controlled by this methodology.

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Semi-Automatic Method for Early Detection of Xylella fastidiosa in Olive Trees Using UAV Multispectral Imagery and Geostatistical-Discriminant Analysis

Remote Sensing ◽

10.3390/rs13010014 ◽

2020 ◽

Vol 13 (1) ◽

pp. 14

Author(s):

Annamaria Castrignanò ◽

Antonella Belmonte ◽

Ilaria Antelmi ◽

Ruggiero Quarto ◽

Francesco Quarto ◽

...

Keyword(s):

Discriminant Analysis ◽

Early Detection ◽

Plant Pathogens ◽

Early Stage ◽

Xylella Fastidiosa ◽

Mediterranean Countries ◽

Parametric Classification ◽

Aerial Vehicle ◽

Olive Groves ◽

Olive Trees

Xylella fastidiosa subsp. pauca (Xfp) is one of the most dangerous plant pathogens in the world. Identified in 2013 in olive trees in south–eastern Italy, it is spreading to the Mediterranean countries. The bacterium is transmitted by insects that feed on sap, and causes rapid wilting in olive trees. The paper explores the use of Unmanned Aerial Vehicle (UAV) in combination with a multispectral radiometer for early detection of infection. The study was carried out in three olive groves in the Apulia region (Italy) and involved four drone flights from 2017 to 2019. To classify Xfp severity level in olive trees at an early stage, a combined method of geostatistics and discriminant analysis was implemented. The results of cross-validation for the non-parametric classification method were of overall accuracy = 0.69, mean error rate = 0.31, and for the early detection class of accuracy 0.77 and misclassification probability 0.23. The results are promising and encourage the application of UAV technology for the early detection of Xfp infection.

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HPLC-MS/MS method applied to an untargeted metabolomics approach for the diagnosis of “olive quick decline syndrome”

Analytical and Bioanalytical Chemistry ◽

10.1007/s00216-021-03279-7 ◽

2021 ◽

Author(s):

Sabrina Di Masi ◽

Giuseppe E. De Benedetto ◽

Cosimino Malitesta ◽

Maria Saponari ◽

Cinzia Citti ◽

...

Keyword(s):

Bacterial Infections ◽

High Performance ◽

High Resolution Mass Spectrometry ◽

Xylella Fastidiosa ◽

Long Chain Fatty Acids ◽

Powerful Approach ◽

Diffusible Signal Factor ◽

Profile Changes ◽

Olive Quick Decline Syndrome ◽

Olive Trees

AbstractOlive quick decline syndrome (OQDS) is a disorder associated with bacterial infections caused by Xylella fastidiosa subsp. pauca ST53 in olive trees. Metabolic profile changes occurring in infected olive trees are still poorly investigated, but have the potential to unravel reliable biomarkers to be exploited for early diagnosis of infections. In this study, an untargeted metabolomic method using high-performance liquid chromatography coupled to quadrupole-time-of-flight high-resolution mass spectrometry (HPLC-ESI-Q-TOF-MS) was used to detect differences in samples (leaves) from healthy (Ctrl) and infected (Xf) olive trees. Both unsupervised and supervised data analysis clearly differentiated the groups. Different metabolites have been identified as potential specific biomarkers, and their characterization strongly suggests that metabolism of flavonoids and long-chain fatty acids is perturbed in Xf samples. In particular, a decrease in the defence capabilities of the host after Xf infection is proposed because of a significant dysregulation of some metabolites belonging to flavonoid family. Moreover, oleic acid is confirmed as a putative diffusible signal factor (DSF). This study provides new insights into the host-pathogen interactions and confirms LC-HRMS-based metabolomics as a powerful approach for disease-associated biomarkers discovery in plants. Graphical abstract

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Controlling the Spatial Spread of a Xylella Epidemic

Bulletin of Mathematical Biology ◽

10.1007/s11538-021-00861-z ◽

2021 ◽

Vol 83 (4) ◽

Author(s):

Sebastian Aniţa ◽

Vincenzo Capasso ◽

Simone Scacchi

Keyword(s):

Spatial Structure ◽

Numerical Simulations ◽

Control Strategies ◽

Xylella Fastidiosa ◽

Cost Effective ◽

Olive Tree ◽

Control Measures ◽

Weed Biomass ◽

Spatial Spread ◽

Olive Trees

AbstractIn a recent paper by one of the authors and collaborators, motivated by the Olive Quick Decline Syndrome (OQDS) outbreak, which has been ongoing in Southern Italy since 2013, a simple epidemiological model describing this epidemic was presented. Beside the bacterium Xylella fastidiosa, the main players considered in the model are its insect vectors, Philaenus spumarius, and the host plants (olive trees and weeds) of the insects and of the bacterium. The model was based on a system of ordinary differential equations, the analysis of which provided interesting results about possible equilibria of the epidemic system and guidelines for its numerical simulations. Although the model presented there was mathematically rather simplified, its analysis has highlighted threshold parameters that could be the target of control strategies within an integrated pest management framework, not requiring the removal of the productive resource represented by the olive trees. Indeed, numerical simulations support the outcomes of the mathematical analysis, according to which the removal of a suitable amount of weed biomass (reservoir of Xylella fastidiosa) from olive orchards and surrounding areas resulted in the most efficient strategy to control the spread of the OQDS. In addition, as expected, the adoption of more resistant olive tree cultivars has been shown to be a good strategy, though less cost-effective, in controlling the pathogen. In this paper for a more realistic description and a clearer interpretation of the proposed control measures, a spatial structure of the epidemic system has been included, but, in order to keep mathematical technicalities to a minimum, only two players have been described in a dynamical way, trees and insects, while the weed biomass is taken to be a given quantity. The control measures have been introduced only on a subregion of the whole habitat, in order to contain costs of intervention. We show that such a practice can lead to the eradication of an epidemic outbreak. Numerical simulations confirm both the results of the previous paper and the theoretical results of the model with a spatial structure, though subject to regional control only.

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Further In Vitro Assessment and Mid-Term Evaluation of Control Strategy of Xylella fastidiosa subsp. pauca in Olive Groves of Salento (Apulia, Italy)

Pathogens ◽

10.3390/pathogens10010085 ◽

2021 ◽

Vol 10 (1) ◽

pp. 85

Author(s):

Giuseppe Tatulli ◽

Vanessa Modesti ◽

Nicoletta Pucci ◽

Valeria Scala ◽

Alessia L’Aurora ◽

...

Keyword(s):

Control Strategy ◽

Xylella Fastidiosa ◽

In Planta ◽

Bacterial Concentration ◽

Infected Area ◽

Olive Groves ◽

Term Evaluation ◽

In Vitro Assessment ◽

Olive Trees

During recent years; Xylella fastidiosa subsp. pauca (Xfp) has spread in Salento causing relevant damage to the olive groves. Measures to contain the spreading of the pathogen include the monitoring of the areas bordering the so-called “infected” zone and the tree eradication in case of positive detection. In order to provide a control strategy aimed to maintain the tree productivity in the infected areas, we further evaluated the in vitro and in planta mid-term effectiveness of a zinc-copper-citric acid biocomplex. The compound showed an in vitro bactericidal activity and inhibited the biofilm formation in representative strains of X. fastidiosa subspecies, including Xfp isolated in Apulia from olive trees. The field mid-term evaluation of the control strategy assessed by quantitative real-time PCR in 41 trees of two olive groves of the “infected” area revealed a low concentration of Xfp over the seasons upon the regular spraying of the biocomplex over 3 or 4 consecutive years. In particular, the bacterial concentration lowered in July and October with respect to March, after six consecutive treatments. The trend was not affected by the cultivar and it was similar either in the Xfp-sensitive cultivars Ogliarola salentina and Cellina di Nardò or in the Xfp-resistant Leccino. Moreover, the scoring of the number of wilted twigs over the seasons confirmed the trend. The efficacy of the treatment in the management of olive groves subjected to a high pathogen pressure is highlighted by the yielded a good oil production

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