Periodic Reporting for period 1 - XYL-SPIT (Understanding the relationship between Xylella fastidiosa and the meadow spittlebug Philaenus spumarius for a sustainable bacterial-mediated diseases control)
Reporting period: 2020-01-16 to 2022-01-15
Research on sustainable containment strategies of diseases caused by the vector-borne bacterium Xylella fastidiosa is hampered by the lack of knowledge on key elements of vector-host-bacterium intimate interaction. Moreover, available data on this three-way relationship come from studies carried out with sharpshooters, the main American bacterial vectors. On the other side, nothing is known on the insect family who likely plays the major role in bacterium epidemiology in Europe, i.e. spittlebugs, to which Philaenus spumarius belongs to. The main objective of the XYL-SPIT project was therefore to thoroughly characterize P. spumarius-X. fastidiosa relationship and transmission dynamics through a multidisciplinary approach, combining real-time tracking of insect probing behavior and classical transmission tests. In addition, we aimed at understanding the role played by vector behavior on X. fastidiosa prevalence on olive varieties with a different degree of susceptibility to the fastidious bacterium observed in the Southern Italian outbreak.
In WP1, the fellow conducted EPG-assisted transmission experiments with both the meadow spittlebug and Graphocephala atropunctata (sharpshooter) that will conclusively identify the feeding behavioral pattern(s) conducive to the bacterium inoculation into the host plant. Molecular analysis on insects and plants, and analysis of the EPG data, are currently ongoing, with final results and a tentative manuscripts expected for summer 2022. In addition, the feeding behavioral comparison between insect species competent (the spittlebug and the sharpshooter) and non-competent (the Issidae Agalmatium curtulum) for X. fastidiosa transmission will furnish indications regarding the acquisition dynamics.
The only substantial change to the original proposal was made for WP2: the original aim of WP2 was the detection of binding/release sites of X. fastidiosa on P. spumarius (the meadow spittlebug) foregut. However, possibly due to the relatively low acquisition efficiency of X. fastidiosa by P. spumarius, none of the 81 samples prepared and analyzed by Scanning Electron Microscopy (SEM) carried bacterial cells. As contingency measure, given also the delay in project implementation due to restrictions associated to Covid pandemic, we opted for focusing on the analysis of the effect of foregut colonization by X. fastidiosa on P. spumarius feeding behavior, metabolism, and immune response. The results that have been produced (see below: published and submitted manuscripts), or are being produced (part of the molecular analysis are still ongoing, with final data expected for end of the spring 2022. Immune response experiments will be carried out during spring 2022), will constitute a significant advancement of the knowledge of spittlebug-Xylella fastidiosa interaction and transmission mechanisms, and could also be exploited for the development of sustainable vector control strategies and tools alternative to pesticides.
Finally, WP3 highlighted the likely fundamental role played by vector behavior, in terms of preference, location, and acceptance, in shaping the disease prevalence on olive varieties (Ogliarola, Leccino, FS-17) with a different degree of susceptibility to X. fastidiosa observed in Southern Italy. Philaenus spumarius tends to significantly prefer the susceptible (Ogliarola) to the tolerant (Leccino and FS-17) varieties when offered a choice. Leccino resulted to be the least favorite among the varieties tested, according to the combined results of EPG and survival tests. We found a strict correlation between the host plant suitability for the spittlebug (expressed in terms of total duration of the xylem sap ingestion) and the concentration of malate in the xylem sap; possibly, the xylem sap ingestion duration is inversely correlated to the concentration of malic acid, and/or with xylem sap pH. On the contrary, differences in xylem vessels anatomy among the olive varieties do not significantly influence spittlebug feeding.
Daniele Cornara carried out and was involved in research activities on vectors of Xylella fastidiosa whose results were published in indexed scientific journals (5 manuscripts). Two manuscripts, one investigating the effects of insecticides on the meadow spittlebug feeding behavior and bacterium transmission (Lago et al.), and the other comparing sharpshooters and spittlebugs feeding behavior and proposing new open-source protocol and software for the analysis of the EPG data (Markheiser et al.), are currently under-review. A reference to EU funding to D. Cornara was always included in the funding/acknowledgement section. The manuscripts have been and will be made available on Daniele Cornara Research gate profile Daniele Cornara (researchgate.net) and on the public repository Zenodo.
In addition, we expect to produce at least three manuscripts resulting directly from the activities carried out in the frame of the XYL-SPIT project, with molecular analysis of insects and plants used in the transmission essays currently ongoing: i) Characterization of the X. fastidiosa inoculation dynamic by sharpshooters and spittlebugs; ii) Evaluation of X. fastidiosa on the meadow spittlebug feeding behavior, metabolism, and immune response; iii) Host selection, acceptance, and feeding behavior of the meadow spittlebug on olive varieties susceptible and resistant to X. fastidiosa.
The researcher is currently planning for summer 2022 a dissemination activity of project results dedicated to researchers, stakeholders and general public in the area of the first Southern Italian X. fastidiosa outbreak along with the project partners, and in collaboration with the University of Bari and CIHEAM-Bari. The XYL-SPIT activities and results have been also disseminated through:
• Interview on an Italian television educational show dedicated to young researchers Vita da Ricercatore 2019 - Daniele Cornara - Video - RaiPlay
• XYL-SPIT project Twitter profile (2) XYL-SPIT - Marie Sklodowska-Curie Fellowship (@XylSpit) / Twitter
• Daniele Cornara Research Gate personal profile Daniele Cornara (researchgate.net)
• Lecture “Electronic monitoring of insects feeding behavior and pathogen transmission”. 03 December 2021 UC Berkeley, Essig’s brunch. Referent: Prof. R. Almeida
• 15th International Symposium of Plant Virus Epidemiology (ISPVE) “Epidemiology and management of plant viruses under a changing climate” 5th-8th June, 2022, Madrid (Spain)
The only substantial change to the original proposal was made for WP2: the original aim of WP2 was the detection of binding/release sites of X. fastidiosa on P. spumarius (the meadow spittlebug) foregut. However, possibly due to the relatively low acquisition efficiency of X. fastidiosa by P. spumarius, none of the 81 samples prepared and analyzed by Scanning Electron Microscopy (SEM) carried bacterial cells. As contingency measure, given also the delay in project implementation due to restrictions associated to Covid pandemic, we opted for focusing on the analysis of the effect of foregut colonization by X. fastidiosa on P. spumarius feeding behavior, metabolism, and immune response. The results that have been produced (see below: published and submitted manuscripts), or are being produced (part of the molecular analysis are still ongoing, with final data expected for end of the spring 2022. Immune response experiments will be carried out during spring 2022), will constitute a significant advancement of the knowledge of spittlebug-Xylella fastidiosa interaction and transmission mechanisms, and could also be exploited for the development of sustainable vector control strategies and tools alternative to pesticides.
Finally, WP3 highlighted the likely fundamental role played by vector behavior, in terms of preference, location, and acceptance, in shaping the disease prevalence on olive varieties (Ogliarola, Leccino, FS-17) with a different degree of susceptibility to X. fastidiosa observed in Southern Italy. Philaenus spumarius tends to significantly prefer the susceptible (Ogliarola) to the tolerant (Leccino and FS-17) varieties when offered a choice. Leccino resulted to be the least favorite among the varieties tested, according to the combined results of EPG and survival tests. We found a strict correlation between the host plant suitability for the spittlebug (expressed in terms of total duration of the xylem sap ingestion) and the concentration of malate in the xylem sap; possibly, the xylem sap ingestion duration is inversely correlated to the concentration of malic acid, and/or with xylem sap pH. On the contrary, differences in xylem vessels anatomy among the olive varieties do not significantly influence spittlebug feeding.
Daniele Cornara carried out and was involved in research activities on vectors of Xylella fastidiosa whose results were published in indexed scientific journals (5 manuscripts). Two manuscripts, one investigating the effects of insecticides on the meadow spittlebug feeding behavior and bacterium transmission (Lago et al.), and the other comparing sharpshooters and spittlebugs feeding behavior and proposing new open-source protocol and software for the analysis of the EPG data (Markheiser et al.), are currently under-review. A reference to EU funding to D. Cornara was always included in the funding/acknowledgement section. The manuscripts have been and will be made available on Daniele Cornara Research gate profile Daniele Cornara (researchgate.net) and on the public repository Zenodo.
In addition, we expect to produce at least three manuscripts resulting directly from the activities carried out in the frame of the XYL-SPIT project, with molecular analysis of insects and plants used in the transmission essays currently ongoing: i) Characterization of the X. fastidiosa inoculation dynamic by sharpshooters and spittlebugs; ii) Evaluation of X. fastidiosa on the meadow spittlebug feeding behavior, metabolism, and immune response; iii) Host selection, acceptance, and feeding behavior of the meadow spittlebug on olive varieties susceptible and resistant to X. fastidiosa.
The researcher is currently planning for summer 2022 a dissemination activity of project results dedicated to researchers, stakeholders and general public in the area of the first Southern Italian X. fastidiosa outbreak along with the project partners, and in collaboration with the University of Bari and CIHEAM-Bari. The XYL-SPIT activities and results have been also disseminated through:
• Interview on an Italian television educational show dedicated to young researchers Vita da Ricercatore 2019 - Daniele Cornara - Video - RaiPlay
• XYL-SPIT project Twitter profile (2) XYL-SPIT - Marie Sklodowska-Curie Fellowship (@XylSpit) / Twitter
• Daniele Cornara Research Gate personal profile Daniele Cornara (researchgate.net)
• Lecture “Electronic monitoring of insects feeding behavior and pathogen transmission”. 03 December 2021 UC Berkeley, Essig’s brunch. Referent: Prof. R. Almeida
• 15th International Symposium of Plant Virus Epidemiology (ISPVE) “Epidemiology and management of plant viruses under a changing climate” 5th-8th June, 2022, Madrid (Spain)
Overall, the XYL-SPIT project produced and will produce knowledge pivotal for developing new sustainable tools for an environmentally friendly control of X. fastidiosa-mediated diseases, based on vector-bacterium interaction disruption and the use of varieties with defensive traits.
In addition, The MSC fellowship, and the international collaborations established during it, played a significant role for the fellow for the achievement of a permanent academic position. Indeed, from February 2022, D. Cornara holds a position of Assistant Professor at the University of Bari. The fellow will bring to the new institution the expertise acquired during the fellowship, and will strengthen the international collaboration with the project partners, trough shared research activities, meetings, and by hosting/asking to host to the partners scholars interested in insect behavior associated with the transmission of plant pathogens.
In addition, The MSC fellowship, and the international collaborations established during it, played a significant role for the fellow for the achievement of a permanent academic position. Indeed, from February 2022, D. Cornara holds a position of Assistant Professor at the University of Bari. The fellow will bring to the new institution the expertise acquired during the fellowship, and will strengthen the international collaboration with the project partners, trough shared research activities, meetings, and by hosting/asking to host to the partners scholars interested in insect behavior associated with the transmission of plant pathogens.