(1) Background: Since 2013, the pathogenic bacterium Xylella fastidiosa has been severely affecting olive production in Apulia, Italy, with consequences for the economy, local culture, landscape and biodiversity. The production of a phenolic extract from fresh olive leaves was employed for endotherapeutic injection into naturally infected olive trees by Xylella fastidiosa in Apulia region, Italy. (2) Methods: The effectiveness of the extract was tested in vitro and in planta in comparison with analogous treatments based on garlic powder and potassium phosphite. (3) Results: The uptake of phenolic compounds from olive leaves through a trunk injection system device resulted in a statistically significant increase in leaf area index and leaf area density, as well as in the growth of newly formed healthy shoots. Plant growth-promoting effects were also observed for potassium phosphite. Moreover, the bacteriostatic activities of the phenolic extract and of the garlic-powder-based solution have been demonstrated in in vitro tests. (4) Conclusions: The results obtained and the contained costs of extraction make the endotherapeutic treatment with phenolic compounds a promising strategy for controlling X fastidiosa to be tested on a larger scale, although the experiments conducted in this study proved not to be suitable for centenary trees.

The meadow spittlebug Philaenus spumarius (Hemiptera: Aphrophoridae) is distributed in several habitats worldwide and has been recently recognized as the main vector of Xylella fastidiosa subsp. pauca. This bacterium has been associated with olive quick decline syndrome (OQDS) in the Salento Peninsula (Italy) and is responsible for extensive desiccation and die-off of olive trees. Current OQDS management strategies include the control of P. spumarius populations, mainly through the removal of weed hosts and insecticide treatments. In addition to the mandatory phytosanitary measures, the demand for new strategies compatible with integrated pest management is increasing. In this study, laboratory biological assays were performed to assess the potential toxic effect of vegetal formulations against P. spumarius adults. Two formulations were tested at different concentrations: Form A, an emulsion of 10% hot pepper-infused oil (Capsicum annuum subspecies Cayenna in olive oil) and Arabic gum in an aqueous solution of extracts of Salvia guaranitica, and Form B, an aqueous solution of extracts of Taxus baccata. Both Form A and B showed to be toxic against P. spumarius compared to the water control. The mean percentage of spittlebug mortality obtained with Form A one hour after treatments was dose–dependent; the lethal dose values were 0.13% (LD25), 0.36% (LD50), and 0.85% (LD75). At the same time, no significant differences in mortality rate were observed between the 0.75% treatment and the treatments with deltamethrin (about 90%). The mean percentage of spittlebug mortality obtained with Form B ranged from 21% to 53% one hour after treatment, but these values were significantly lower than those obtained with deltamethrin. The effectiveness of Form A on the P. spumarius population was also evaluated in the field. The averages of captures in the three experimental blocks were 1.8/trap for treated and 7.7/trap for untreated plots, and the spittlebug populations significantly decreased after treatments. Based on these results and the literature data, we hypothesize that the effectiveness of Form A is the result of the synergistic effect of all its components. No symptoms of phytotoxicity were recorded on olive trees treated with Form A, and the number of P. spumarius specimens collected on these plants was much lower than on untreated plants. These results suggest the potential use of Form A in the protection of olive trees. This vegetal formulation can thus be considered as a valid alternative to chemical insecticides for the control of the main vector of X. fastidiosa and could be integrated into a sustainable management system for OQDS.

In 2022, a test performance study (TPS) assessing the influence of different master mixes on the performance of the tetraplex real-time PCR (TqPCR) assay was organized. TqPCR allows for the specific detection and identification of Xylella fastidiosa (Xf) subspecies in a single reaction. Eighteen official laboratories of the Italian National Plant Protection Organization received a panel of 12 blind samples, controls, primers, probes, and different master mixes to participate in the TPS. Furthermore, the Research Centre for Plant Protection and Certification of the Council for Agricultural Research and Economics performed an intra-laboratory study (ITS) on spiked plant matrices to evaluate the analytical sensitivity of TqPCR employing the selected master mixes with the best performance. Naturally infected samples were analyzed for subspecies identification via TqPCR compared with the official multilocus-sequence-typing (MLST) method. The best results in this comparative study were obtained using Fast Universal PCR Master Mix (Applied Biosystems) and Brilliant multiplex QPCR Master Mix (Agilent), and they confirmed that the TqPCR test is reliable, offering the advantage of identifying this subspecies at the same time, thus saving time and resources. The TqPCR assay is suggested among the tests to be used by laboratories performing the official diagnosis of Xf to support the activities of official monitoring.

The bacterial pathogen Xylella fastidiosa (Xf), which causes several plant diseases with significant economic impacts on agriculture and the environment, remains a challenge to manage due to its wide host range. This study investigated the in vitro antibacterial effects of natural compounds, including Trametes versicolor extract, clove essential oil, and the resistance inducer FossilⓇ, against X. fastidiosa subsp. fastidiosa using an antibacterial susceptibility testing (AST) method based on microfluidic channels. This novel method was compared with the traditional broth macrodilution method to assess its reliability and the potential advantages microfluidics offers. For each substance and test, both the ability to limit planktonic growth (reported as the minimum inhibitory concentration) and the ability to inhibit biofilm formation were evaluated. The results suggest that compared to the macrodilution method, microfluidic channels allow for a more rapid AST execution, use less material, and allow for real-time observation of bacterial behavior under a continuous flow of nutrients and antibacterial substances. All tested products demonstrated high antibacterial efficacy against Xf with the macrodilution method, yielding comparable results with microfluidic AST. These findings highlight the antimicrobial properties of the tested substances and establish the groundwork for applying this new technique to select promising eco-friendly products for potential future field applications in controlling Xf.