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Behavioral manipulation (BM) is a multimodal control approach based on the interference with the stimuli mediating insect perception and interaction with the surroundings. BM could represent a win–win strategy for the management of vector-borne plant pathogens as the bacterium Xylella fastidiosa, since it could reduce the number of vectors alighting on host plants and, consequently, the chances for transmission to occur. In this review, we summarized current knowledge and highlighted gaps in information on (i) how insect vectors of X. fastidiosa in general, and more specifically the meadow spittlebug Philaenus spumarius, locate and accept the host plant; and (ii) how behavioral manipulation techniques could be applied to disrupt the vector–host plant interaction. Finally, we discussed how diverse BM strategies could be combined with other integrated pest management tools to protect olive groves from inoculation with the fastidious bacterium.

BACKGROUND Many groups of insects utilize substrate-borne vibrations for intraspecific communication. This characteristic makes them a suitable model for exploring the use of vibrations as a tool for pest control as an alternative to the use of chemicals. Detailed knowledge of species communication is a prerequisite to select the best signals to use. This study explored the use of substrate-borne vibrations for pest control of the brown marmorated stink bug (BMSB), Halyomorpha halys Stal (Heteroptera: Pentatomidae). For this purpose, we first identified the spectral and temporal characteristics that best elicit male responsiveness. Bioassays were conducted with artificial signals that mimicked the natural female calling signal. Second, we used the acquired knowledge to synthesize new signals endowed with different degrees of attractiveness in single- and two-choice bioassays using a wooden custom-made T stand. RESULTS The results from this study showed that males were attracted to female signals along a high range of amplitudes, especially starting from a threshold of 100 μm s-1 , a high pulse repetition time (1 s) and frequency peak corresponding to the first harmonic (76 Hz). This resulted in an "optimal" signal for use to attract males, while the choice test in the T arena showed that this signal elicits searching behavior and attracts BMSB males towards a stimulation point. CONCLUSION We confirm the use of vibrational signals as a strong tool for behavioral manipulation of male BMSB and suggest its possible use in the development of field traps and further management of this pest. © 2021 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Insecticides are essential, though controversial tools in modern pest management. Insecticides can slow the spread of key vector-borne plant pathogens, but often lead to inconsistent results given that insecticide use is generally focused on acute toxicity under no-choice conditions. Here, we analysed the lethal (survival) and sublethal (feeding behaviour) effects of six commercial products (acetamiprid, deltamethrin, spinosad, sulfoxaflor, pyrethrin and kaolin) on Philaenus spumarius, vector of the bacterium Xylella fastidiosa. Furthermore, we assessed the impact of insecticides displaying different degrees of acute toxicity against spittlebugs (highest to lowest: acetamiprid, pyrethrin and kaolin) on the transmission of X. fastidiosa by P. spumarius under both free-choice and no-choice conditions.Deltamethrin, acetamiprid and to a limited extent pyrethrin significantly altered the feeding behaviour of P. spumarius. Deltamethrin and acetamiprid were highly toxic against P. spumarius, but the mortality induced by exposure to pyrethrin was limited overall. By contrast, spinosad, sulfoxaflor and kaolin did not significantly impact P. spumarius feeding behaviour or survival. Under no-choice conditions, both pyrethrin and acetamiprid reduced the X. fastidiosa inoculation rate compared with kaolin and the control. On the other hand, pyrethrin reduced transmission, but acetamiprid failed to significantly affect bacterial inoculation under free-choice conditions.Pyrethrin was the only compound able to reduce X. fastidiosa transmission under both free-choice and no-choice conditions. Xylella fastidiosa management strategy based exclusively on the evaluation of insecticide acute toxicity under no-choice conditions would most likely fail to prevent, or slow, bacterial spread. © 2022 The Authors. Pest Management Science published by John WileySons Ltd on behalf of Society of Chemical Industry.

The bacterium Xylella fastidiosa (Xf) is an invasive insect-borne pathogen, which causes lethal diseases to important crops including olives, citrus, almonds and grapes as well as numerous forest, ornamental, and uncultivated plants. Outbreaks of Xf-related plant diseases are currently occurring in the Mediterranean region, causing substantial losses to various agricultural sectors. Several models have recently been published to identify which regions are at highest risk in Europe; however, such models did not consider the insect vectors, which constitute the key driver of short-range Xf spread. We fitted bioclimatic species distribution models to depict the macroclimatic preferences of the meadow spittlebug Philaenus spumarius L. (1978) (Hemiptera: Aphrophoridae), the major epidemiologically relevant vector currently responsible for Xf spread in the Europe. Many regions of Western Europe and Mediterranean basin are predicted by models as highly climatically suitable for this vector, including all regions where severe Xf have occurred so far. Conversely, the driest and warmest areas of the Mediterranean basin are predicted as little suitable for P. spumarius. Models forecast that agricultural-important parts of the southern Mediterranean area might experience a substantial decrease in climatic suitability for P. spumarius by the period 2040–2060. Areas predicted as highly suitable just for the bacterium but not optimal for this vector are apparently still free of severe Xf outbreaks, suggesting that climate tolerances of P. spumarius might partly explain the current spatial pattern of Xf outbreaks in Europe and should always be considered in further risk assessments.

Scaphoideus titanus (Ball) is a grapevine-feeder leafhopper, and the most important vector of Flavescence dorée of grapevine (FD), a disease associated with phytoplasmas belonging to ribosomal subgroups 16Sr-V-C and -D. FD is a major constraint to viticulture in several European countries and, so far, its control has relied on roguing of infected plants and insecticide applications against the vector. Detailed knowledge on different levels of the multifaceted phytoplasma-plant-vector relationship is required to envisage and explore more sustainable ways to control the disease spread. In the present work, S. titanus feeding behaviour was described on three grapevine cultivars: Barbera (susceptible to FD), Brachetto, and Moscato (tolerant to FD) using the Electrical Penetration Graph (EPG) technique. Interestingly, no differences were highlighted in the non-phloem feeding phases, thus suggesting that the tested cultivars have no major differences in the biochemical composition or structure of the leaf cuticle, epidermis or mesophyll, that can affect the first feeding activities. On the contrary, the results showed significant differences in leafhopper feeding behaviour in terms of the duration of the phloem feeding phase, longer on Barbera and shorter on Brachetto and Moscato, and of the frequency of interruption-salivation events inside the phloem, higher on Brachetto and Moscato. These findings indicate a possible preference for the Barbera cultivar, a better host for the leafhopper. Scaphoideus titanus feeding behaviour on Barbera correlates with an enhanced FDp transmission efficiency, thus explaining, at least in part, the higher susceptibility of this cultivar to FD. The mechanisms for the possible non-preference for Brachetto and Moscato are discussed, and an antixenosis is hypothesized. We propose that breeding for resistance against FD should take into account both plant traits associated with the response to the phytoplasmas and to the vector.

Scaphoideus titanus (Ball) is a grapevine-feeder leafhopper, and the most important vector of Flavescence dorée of grapevine (FD), a disease associated with phytoplasmas belonging to ribosomal subgroups 16Sr-V–C and –D. FD is a major constraint to viticulture in several European countries and, so far, its control has relied on roguing of infected plants and insecticide applications against the vector. Detailed knowledge on different levels of the multifaceted phytoplasma-plant-vector relationship is required to envisage and explore more sustainable ways to control the disease spread. In the present work, S. titanus feeding behaviour was described on three grapevine cultivars: Barbera (susceptible to FD), Brachetto, and Moscato (tolerant to FD) using the Electrical Penetration Graph (EPG) technique. Interestingly, no differences were highlighted in the non-phloem feeding phases, thus suggesting that the tested cultivars have no major differences in the biochemical composition or structure of the leaf cuticle, epidermis or mesophyll, that can affect the first feeding activities. On the contrary, the results showed significant differences in leafhopper feeding behaviour in terms of the duration of the phloem feeding phase, longer on Barbera and shorter on Brachetto and Moscato, and of the frequency of interruption-salivation events inside the phloem, higher on Brachetto and Moscato. These findings indicate a possible preference for the Barbera cultivar, a better host for the leafhopper. Scaphoideus titanus feeding behaviour on Barbera correlates with an enhanced FDp transmission efficiency, thus explaining, at least in part, the higher susceptibility of this cultivar to FD. The mechanisms for the possible non-preference for Brachetto and Moscato are discussed, and an antixenosis is hypothesized. We propose that breeding for resistance against FD should take into account both plant traits associated with the response to the phytoplasmas and to the vector.

Philaenus spumarius was identified as the predominant vector involved in the spread of Xylella fastidiosa in Europe. Studies on pesticides lethal and sub-lethal effects on insect vectors are essential for the development of sustainable control strategies. Therefore, we studied the effect of six pesticides (acetamiprid, delthametrin, spinosad, sulfoxaflor, pyrethrin, kaolin) on the survival and feeding behavior of P. spumarius when exposed to treated plants (Sonchus olearaceous) through the Electrical Penetration Graph (EPG) technique. First we observed the spittlebug feeding behaviour for 4h on treated plants; second, insect survival was assessed for 4h on treated plants and the feeding behavior of survived individuals was studied on untreated plants. In addition, we evaluated the effect of pyrethrin, acetamiprid, and kaolin on the transmission of X. fastidiosa ST53 to Catharanthus roseus plants. We carried out “no-choice” transmission tests where insects were caged on plants previously treated with a single compound or “free choice” trials with spittlebugs confined in a cage containing plants treated with the 3 compounds. We found that plants treated with deltamethrin and acetamiprid had a repellent effect (insects escaped) compared to other treatments. These pesticides reduced the duration of Xc (xylem contact), Xi (xylem ingestion) and the time to the first sustained Xi, when compared to control and kaolin. In the second assay, all insects exposed to deltamethrin or acetamiprid died in two hours. No significant differences in the feeding behavior of insects on untreated plants after the exposure to the tested compounds were found. Pyrethrin and acetamiprid reduced the transmission rate of X. fastidiosa by P. spumarius while kaolin gave similar results as the control untreated plants. Acetamiprid, however failed to reduce the transmission rate under free choice conditions. Overall, our results suggest that pyrethrin and deltamethrin are the most promising compounds to manage the transmission of X. fastidiosa.

The Orange Spiny Whitefly (OSW) Aleurocanthus spiniferus (Hemiptera: Aleyrodidae) represents a new serious threat to Citrus spp., grapevine and ornamental plants in the whole Mediterranean area. Such threat urgently calls for the development of a sustainable control strategy, including insecticides compatible with biological control, and applicable also in organic citrus farming that represent an essential part of Mediterranean agricultural economy. Therefore, we evaluated the toxicity and the effects on host searching, oviposition, and probing and feeding behavior exerted on OSW by organic insecticides supposed to have limited side effects on environment and ecosystem services, i.e. sweet orange essential oil (EO), extract of Clitoria ternatea (CT), mineral oil, pyrethrin and azadirachtin. Despite none of the compounds caused a significant mortality of any of the OSW instars, we observed interesting effects on whitefly behavior: (i) EO and pyrethrin showed a relevant repellent effect, with impairment of both adults landing and oviposition on treated plants; (ii) CT and pyrethrin strongly affected probing behavior. Here, in the light of our findings, we discuss possible OSW sustainable control strategies and further research perspectives.

Cicadas are prominent insects in the Mediterranean basin environments, including in olive orchards. The bacterium Xylella fastidiosa is a xylem-limited vector-borne plant pathogen that was introduced into southern Italy, causing significant losses of olive trees. Cicadas are xylem-sap feeders and potential vectors of X. fastidiosa; however, knowledge of their role in the transmission of this bacterium is limited. We carried out two separate experiments: the first in California (USA), where we verified the capability of Platypedia minor to transmit X. fastidiosa from infected to healthy grapevines; the second in Apulia (South Italy), where we evaluated if Cicada orni collected on infected olive plants were able to transmit the bacterium to healthy olives. In California, transmission was not achieved for the 19 grapevines caged each with a group of two to three P. minor; moreover, none of the 19 insects (out of the 47 used for the transmission test) tested by culturing resulted positive for X. fastidiosa. In Italy, none of the olive recipient plants either caged with groups of three C. orni individuals per plant confined in sleeve cages (55 plants) or placed inside a mesocosm with cicadas free to move among the recipient plants (30 plants), were infected with the bacterium. Moreover, out of the 314 field collected C. orni tested by qPCR, only 4 (1.27%) were positive for the bacterium. Our data suggest that the cicada species we tested likely have no or a negligible role in the natural spread of X. fastidiosa.

The phytosanitary emergence triggered by Xylella fastidiosa introduction in Europe urgently calls for research on its main vector, the meadow spittlebug, Philaenus spumarius. The difficulties faced in altering P. spumarius life cycle setting up a continuous indoor rearing under artificial conditions, obtaining a continuous availability of insects for laboratory trials, represent a great limit for research. Here, we propose a methodology to rear P. spumarius in the laboratory allowing the supply of nymphs and adults before they become available in the field. This crucial step forward will permit to overcome the seasonality barrier, reducing time and efforts currently required for experimentation on the meadow spittlebug. The proposed methodology would allow producing the data urgently required to fill the knowledge gap and finally set up an effective and environmentally friendly control strategy of P. spumarius.

© 2016 The American Phytopathological Society. There is little information available on Xylella fastidiosa transmission by spittlebugs (Hemiptera, Cercopoidea). This group of insect vectors may be of epidemiological relevance in certain diseases, so it is important to better understand the basic parameters of X. fastidiosa transmission by spittlebugs. We used grapevines as a host plant and the aphrophorid Philaenus spumarius as a vector to estimate the effect of plant access time on X. fastidiosa transmission to plants; in addition, bacterial population estimates in the heads of vectors were determined and correlated with plant infection status. Results show that transmission efficiency of X. fastidiosa by P. spumarius increased with plant access time, similarly to insect vectors in another family (Hemiptera, Cicadellidae). Furthermore, a positive correlation between pathogen populations in P. spumarius and transmission to plants was observed. Bacterial populations in insects were one to two orders of magnitude lower than those observed in leafhopper vectors, and population size peaked within 3 days of plant access period. These results suggest that P. spumarius has either a limited number of sites in the foregut that may be colonized, or that fluid dynamics in the mouthparts of these insects is different from that in leafhoppers. Altogether our results indicate that X. fastidiosa transmission by spittlebugs is similar to that by leafhoppers. In addition, the relationship between cell numbers in vectors and plant infection may have under-appreciated consequences to pathogen spread.

Diseases associated with Xylella fastidiosa have been described mostly in North and South America. However, during the last five years, widespread X. fastidiosa infections have been reported in a constrained area of the Apulia region (southern Italy), in olives trees suffering a severe disease, denoted as Olive Quick Decline Syndrome (OQDS). Because many xylem sap-feeding insects can function as vectors for the transmission of this exotic pathogen in EU, several research programs are ongoing to assess the role of candidate vectors in the spread of the infections. Initial investigations identified Philaenus spumarius (L.) as the predominant vector species in the olive orchards affected by the OQDS. Additional experiments have been carried out during 2016 and 2017 to assess the role of other species. More specifically, adults of the spittlebugs Philaenus italosignus Drosopolous and Remane, Neophilaenus campestris (Fallen) and of the planthopper Latilica tunetana (Matsumura) (Issidae) have been tested in transmission experiments to assess their ability to acquire the bacterium from infected olives and to infect different susceptible hosts (olives, almond, myrtle –leaf milkwort, periwinkle). Acquisition rates determined by testing individual insects in quantitative PCR assays, ranging from 5.6% in N. campestris to 22.2% in P. italosignus, whereas no acquisition was recorded for L. tunetana. Successful transmissions were detected in the recipient plants exposed to P. italosignus and N. campestris, whereas no trasmissions occurred with L. tunetana. The known vector Philaenus spumarius has been included in all the experiments for validation. The systematic surveys conducted in 2016 and 2017 provided further evidence on the population dynamics and seasonal abundance of the spittlebug populations in the olive groves.

Artificial diets represent an essential tool for investigations on the intimate relationship between plant pathogens and their vectors. Previous research failed in devising an artificial diet delivery system for the meadow spittlebug Philaenus spumarius, to date considered the most important vector of the bacterium Xylella fastidiosa in Europe. Here, we describe a new delivery “tube system” by which we succeeded in artificial feeding of P. spumarius with holidic diets (one sucrose diet and two amino acids diets). Spittlebug probing and feeding behaviour on either the tube system or a traditional “flat system” realized out of a small Petri dish filled with diet and covered with stretched Parafilm® were observed in real time by video‐EPG (Electrical Penetration Graph), in order to assess the occurrence of ingestion and excretion. Moreover, we evaluated P. spumarius survival on either the tube system filled with the two holidic diets that gave the best EPG results or an empty tube system serving as control. Contrary to the flat system, where just brief stylet insertions through the Parafilm® were recorded, the spittlebug ingested the artificial diets when delivered with the tube system. Survival on the diets provided with the tube system was significantly greater than the control, with no differences between the diets tested. Furthermore, the tube system was suitable also for another spittlebug species shown to be a competent vector of X. fastidiosa, that is Neophilaenus campestris. The tool we devised opens new perspectives for investigations on X. fastidiosa/spittlebugs interactions, as well as for the functional analysis of mutant X. fastidiosa strains in respect to insect colonization and transmission.

Diseases associated with Xylella fastidiosa have been described mostly in North and South America. However, during the last five years, widespread X. fastidiosa infections have been reported in a constrained area of the Apulia region (southern Italy), in olives trees suffering a severe disease, denoted as Olive Quick Decline Syndrome (OQDS). Because many xylem sap-feeding insects can function as vectors for the transmission of this exotic pathogen in EU, several research programs are ongoing to assess the role of candidate vectors in the spread of the infections. Initial investigations identified Philaenus spumarius (L.) as the predominant vector species in the olive orchards affected by the OQDS. Additional experiments have been carried out during 2016 and 2017 to assess the role of other species. More specifically, adults of the spittlebugs Philaenus italosignus Drosopolous and Remane, Neophilaenus campestris (Fallen) and of the planthopper Latilica tunetana (Matsumura) (Issidae) have been tested in transmission experiments to assess their ability to acquire the bacterium from infected olives and to infect different susceptible hosts (olives, almond, myrtle &ndash, leaf milkwort, periwinkle). Acquisition rates determined by testing individual insects in quantitative PCR assays, ranging from 5.6% in N. campestris to 22.2% in P. italosignus, whereas no acquisition was recorded for L. tunetana. Successful transmissions were detected in the recipient plants exposed to P. italosignus and N. campestris, whereas no trasmissions occurred with L. tunetana. The known vector Philaenus spumarius has been included in all the experiments for validation. The systematic surveys conducted in 2016 and 2017 provided further evidence on the population dynamics and seasonal abundance of the spittlebug populations in the olive groves.

The unique color pattern polymorphism and the foamy nymphal case of the meadow spittlebug Philaenus spumarius have attracted the attention of scientists for centuries. Nevertheless, since this species has never been considered a major threat to agriculture, biological, ecological and ethological data are missing and rather scattered. To date this knowledge has become of paramount importance, in view of the discovery of P. spumarius main role in the transmission of the bacterium Xylella fastidiosa in Italy, and possibly in other European countries. The aim of this review is to provide a state of the art about this species, with particular focus on those elements that could help developing environmental friendly and sustainable control programs to prevent transmission of X. fastidiosa. Moreover, recent findings on the role of the meadow spittlebug as vector of the fastidious bacterium within the first reported European bacterium outbreak in Apulia (South Italy) will be discussed.

The bacterium Xylella fastidiosa (Xf) is an invasive insect-borne pathogen, which causes lethal diseases to important crops including olives, citrus, almonds and grapes as well as numerous forest, ornamental, and uncultivated plants. Outbreaks of Xf-related plant diseases are currently occurring in the Mediterranean region, causing substantial losses to various agricultural sectors. Several models have recently been published to identify which regions are at highest risk in Europe; however, such models did not consider the insect vectors, which constitute the key driver of short-range Xf spread. We fitted bioclimatic species distribution models to depict the macroclimatic preferences of the meadow spittlebug Philaenus spumarius L. (1978) (Hemiptera: Aphrophoridae), the major epidemiologically relevant vector currently responsible for Xf spread in the Europe. Many regions of Western Europe and Mediterranean basin are predicted by models as highly climatically suitable for this vector, including all regions where severe Xf have occurred so far. Conversely, the driest and warmest areas of the Mediterranean basin are predicted as little suitable for P. spumarius. Models forecast that agricultural-important parts of the southern Mediterranean area might experience a substantial decrease in climatic suitability for P. spumarius by the period 2040–2060. Areas predicted as highly suitable just for the bacterium but not optimal for this vector are apparently still free of severe Xf outbreaks, suggesting that climate tolerances of P. spumarius might partly explain the current spatial pattern of Xf outbreaks in Europe and should always be considered in further risk assessments. Open Access funding provided thanks to the CRUE-CSIC agreement with Springer Nature. This work has been financially supported by the Spanish Ministerio de Ciencia, Innovación y Universidades, Grant/Award Number: AGL2017-89604-R and the European Union Horizon 2020 research and innovation program under grant agreements no. 635646 POnTE (Pest Organisms Threatening Europe), and no. 727987 XF-ACTORS (Xylella Fastidiosa Active Containment Through a multidisciplinary-Oriented Research Strategy). The first author of this study was funded by the fellowship “Ayudas destinadas a la atracción de talento investigador de la Comunidad de Madrid”. Daniele Cornara participation in this work was supported by a research grant in the frame of European Union’s Horizon 2020 research and innovation programme under the Marie Skłodowska-Curie grant agreement No 835732 XYL-SPIT. info:eu-repo/semantics/publishedVersion

Discovery of Xylella fastidiosa from olive trees with "Olive quick decline syndrome" in October 2013 on the west coast of the Salento Peninsula prompted an immediate search for insect vectors of the bacterium. The dominant xylem-fluid feeding hemipteran collected in olive orchards during a 3-mo survey was the meadow spittlebug, Philaenus spumarius (L.) (Hemiptera: Aphrophoridae). Adult P. spumarius, collected in November 2013 from ground vegetation in X. fastidiosa-infected olive orchards, were 67% (40 out of 60) positive for X. fastidiosa by polymerase chain reaction (PCR) assays. Euscelis lineolatus Brulle were also collected but tested negative for the pathogen. Transmission tests with P. spumarius collected from the Salento area were, therefore, conducted. After a 96-h inoculation access period with 8 to 10 insects per plant and a 30-d incubation period, PCR results showed P. spumarius transmitted X. fastidiosa to two of five periwinkle plants but not to the seven olive plants. Sequences of PCR products from infected periwinkle were identical with those from X. fastidiosa-infected field trees. These data showed P. spumarius as a vector of X. fastidiosa strain infecting olives trees in the Salento Peninsula, Italy.

Introduction: Aleurocanthus spiniferus (Quaintance) is one further alien invasive pest that have been invading the south of Italy. From the EPPO first report of the presence of A. spiniferus, in April 2008, Puglia was invaded northward by this polyphagous whitefly and new territories were infested more or less severely.Aims: This poster refers about recently occupied areas detailing local and urban outbreaks and population density per host plant species. Materials and Methods: We use direct observation on the field and digital camera-equipped stereoscope and Cryo-SEM on the laboratory for data confirmation and imaging. Results: We detail the invaded areas, confirming the spreading of this pest in almost all the Puglia region. Conclusions: The overall ecological preferences and behaviour of the pest are depicted in view of its control in urban areas.

The presence of Aleurocanthusspiniferusis reported in Apulia from April 2008. This polyphagous whitefly is now spreading northward and invading new territories.This paper refers about recently occupied areas detailing local and urban outbreaks and population density per host plant species.We use digital camera-equipped stereoscope and Cryo-SEM on the laboratory and direct observation on the field.We note the presence of A. Spiniferus in Puglia region and we depict the invasion of agricultural and urban areas.The overall behaviour and ecological preferences of the alien pest are discussed in view of its available means of control.

Discovery of Xylella fastidiosa from olive trees with "Olive quick decline syndrome" in October 2013 on the west coast of the Salento Peninsula prompted an immediate search for insect vectors of the bacterium. The dominant xylem-fluid feeding hemipteran collected in olive orchards during a 3-mo survey was the meadow spittlebug, Philaenus spumarius (L.) (Hemiptera: Aphrophoridae). Adult P. spumarius, collected in November 2013 from ground vegetation in X. fastidiosa-infected olive orchards, were 67% (40 out of 60) positive for X. fastidiosa by polymerase chain reaction (PCR) assays. Euscelis lineolatus Brullé were also collected but tested negative for the pathogen. Transmission tests with P. spumarius collected from the Salento area were, therefore, conducted. After a 96-h inoculation access period with 8 to 10 insects per plant and a 30-d incubation period, PCR results showed P. spumarius transmitted X. fastidiosa to two of five periwinkle plants but not to the seven olive plants. Sequences of PCR products from infected periwinkle were identical with those from X. fastidiosa-infected field trees. These data showed P. spumarius as a vector of X. fastidiosa strain infecting olives trees in the Salento Peninsula, Italy.

© 2016, The Author(s). The recent introduction of Xylella fastidiosa in Europe and its involvement in the Olive Quick Decline Syndrome (OQDS) in Apulia (Salento, Lecce district, South Italy) led us to investigate the biology and transmission ability of the meadow spittlebug, Philaenus spumarius, which was recently demonstrated to transmit X. fastidiosa to periwinkle plants. Four xylem-sap-feeding insect species were found within and bordering olive orchards across Salento during a survey carried out from October 2013 to December 2014: P. spumarius was the most abundant species on non-olive vegetation in olive orchards as well as on olive foliage and was the only species that consistently tested positive for the presence of X. fastidiosa using real-time PCR. P. spumarius, whose nymphs develop within spittle on weeds during the spring, are likely to move from weeds beneath olive trees to olive canopy during the dry period (May to October 2014). The first X. fastidiosa-infective P. spumarius were collected in May from olive canopy: all the individuals previously collected on weeds tested negative for the bacterium. Experiments demonstrated that P. spumarius transmitted X. fastidiosa from infected to uninfected olive plants. Moreover, P. spumarius acquired X. fastidiosa from several host plant species in the field, with the highest acquisition rate from olive, polygala and acacia. Scanning electron microscopy (SEM) revealed bacterial cells resembling X. fastidiosa in the foreguts of adult P. spumarius. The data presented here are essential to plan an effective IPM strategy and limit further spread of the fastidious bacterium.

The recent history ofXylella fastidiosaWells et al. introduction in Europe illustrates how the lack of knowledge about the bacterium-vector-host interactions hinders the application of effective containment strategies, with the bacterial spread that currently appears relentless. Vector behavior is a key component in plant pathogen transmission; therefore, detailed knowledge of vector probing behavior on various host plants would furnish useful data for containing the bacterium. Aiming at this goal, we carried out electrical penetration graph (EPG)-assisted probing behavior observations of the spittlebug,Neophilaenus campestris(Fallen) (Hemiptera: Aphrophoridae), a candidate vector widespread in all the areas whereX. fastidiosais currently established. Spittlebug probing was first characterized on one of its preferred host plants,Bromus madritensisL. (Poaceae), over short (6 h) and long (16 h) time spans. Thereafter, we performed comparative observations of the spittlebug probing onB. madritensis, olive, and grapevine. Overall, the probing behavior ofN. campestris, that is, the main characteristic waveforms, did not differ substantially from that ofPhilaenus spumarius(L.) (Hemiptera: Aphrophoridae); however, here we report and describe a new kind of xylem activity interruption, provisionally termed N2, not previously observed inP. spumarius. Considering the time spent in xylem ingestion as a percentage of the total probing time (host-suitability indicator used for spittlebugs), grapevine andB. madritensisare the preferred hosts forN. campestris, with olive as the least suitable among the plants tested. Successful probes, that is, probes during which xylem ingestion occurred, on grapevine were 2.5× greater than olive. Therefore, our data onN. campestrisprobing behavior suggest that the spittlebug role inX. fastidiosaepidemiology in European vineyards should be given more attention. In the present manuscript, we also discuss the current difficulties in interpreting and analyzing the outcomes of EPG studies on spittlebugs.

The Gram-negative bacterium Xylella fastidiosa causes disease in a number of important crops and plants within natural ecosystems. Most notable among these diseases are Pierce’s disease of grapevine, Citrus Variegated Chlorosis, Almond Leaf Scorch, Oleander Leaf Scorch and Olive Quick Decline Syndrome. In response to substantial economic losses, extensive research efforts have been underway over the past several decades in North and South America. Recently, X. fastidiosa has been detected in several countries of Europe and Asia, likely appearing as a result of global trading of plant material. Once established in a new region, X. fastidiosa spread is dependent on the obligate transmission by xylem-sap feeding insects. Sharpshooter leafhoppers (Hemiptera: Cicadellidae: Cicadellinae) are the best-studied group of vectors of this bacterium; however the list of known vector species is expanding. As X. fastidiosa moves into new regions, the combination of an introduced pathogen with existing or introduced vectors carries the potential of increasing economically important disease problems in the local context. By reviewing ecological and biological information of known X. fastidiosa vectors and the epidemiology of local X. fastidiosa outbreaks, this paper aims at increasing our understanding of vector– X. fastidiosa interactions.

Here, we provide the first insights into the transmission dynamics of the bacterium Xylella fastidiosa by the meadow spittlebug Philaenus spumarius, gathered through DC EPG (electrical penetration graph)-assisted transmission tests and comparative observations of the probing and feeding behavior of infective versus non-infective vectors on healthy olive plants. Bacterial cells binding to P. spumarius’ foregut occurred at a very low rate and in a time as short as 15 min spent by the insect in xylem ingestion or activities interspersed with xylem ingestion (interruption during xylem ingestion and resting). P. spumarius inoculation of bacterial cells into the xylem was exclusively associated with an early (ca. 2 to 7 min after the onset of the first probe) and occasional behavior, provisionally termed waveform Xe, presumably related to egestion regulated by pre-cibarial valve fluttering. Infective spittlebugs compared to non-infective ones exhibited: (i) longer non-probing and shorter xylem ingestion; (ii) longer duration of single non-probing events; (iii) fewer sustained ingestions (ingestion longer than 10 min) and interruptions of xylem activity (N); and (iv) longer time required to perform the first probe. These observations suggest difficulties in feeding of infective P. spumarius probably caused by the presence of X. fastidiosa within the foregut. Overall, our data indicate that likely short time—few minutes—is required for X. fastidiosa transmission by P. spumarius; thus, vector control strategies should aim at preventing spittlebug access to the host plant. Furthermore, our findings represent an important contribution for further research on the disruption of spittlebug–bacterium interactions.

Philaenus spumarius was identified as the predominant vector involved in the spread of Xylella fastidiosa in Europe. Studies on pesticides lethal and sub-lethal effects on insect vectors are essential for the development of sustainable control strategies. Therefore, we studied the effect of six pesticides (acetamiprid, delthametrin, spinosad, sulfoxaflor, pyrethrin, kaolin) on the survival and feeding behavior of P. spumarius when exposed to treated plants (Sonchus olearaceous) through the Electrical Penetration Graph (EPG) technique. First we observed the spittlebug feeding behaviour for 4h on treated plants; second, insect survival was assessed for 4h on treated plants and the feeding behavior of survived individuals was studied on untreated plants. In addition, we evaluated the effect of pyrethrin, acetamiprid, and kaolin on the transmission of X. fastidiosa ST53 to Catharanthus roseus plants. We carried out “no-choice” transmission tests where insects were caged on plants previously treated with a single compound or “free choice” trials with spittlebugs confined in a cage containing plants treated with the 3 compounds. We found that plants treated with deltamethrin and acetamiprid had a repellent effect (insects escaped) compared to other treatments. These pesticides reduced the duration of Xc (xylem contact), Xi (xylem ingestion) and the time to the first sustained Xi, when compared to control and kaolin. In the second assay, all insects exposed to deltamethrin or acetamiprid died in two hours. No significant differences in the feeding behavior of insects on untreated plants after the exposure to the tested compounds were found. Pyrethrin and acetamiprid reduced the transmission rate of X. fastidiosa by P. spumarius while kaolin gave similar results as the control untreated plants. Acetamiprid, however failed to reduce the transmission rate under free choice conditions. Overall, our results suggest that pyrethrin and deltamethrin are the most promising compounds to manage the transmission of X. fastidiosa., ES; PPT; clalago.b@gmail.com

Diseases associated with Xylella fastidiosa have been described mostly in North and South America. However, during the last five years, widespread X. fastidiosa infections have been reported in a constrained area of the Apulia region (southern Italy), in olives trees su ering a severe disease, denoted as Olive Quick Decline Syndrome (OQDS). Because many xylem sap-feeding insects can function as vectors for the transmission of this exotic pathogen in EU, several research programs are ongoing to assess the role of candidate vectors in the spread of the infections. Initial investigations identified Philaenus spumarius (L.) as the predominant vector species in the olive orchards a ected by the OQDS. Additional experiments have been carried out during 2016 and 2017 to assess the role of other species. More specifically, adults of the spittlebugs Philaenus italosignus Drosopolous and Remane, Neophilaenus campestris (Fallen) and of the planthopper Latilica tunetana (Matsumura) (Issidae) have been tested in transmission experiments to assess their ability to acquire the bacterium from infected olives and to infect di erent susceptible hosts (olives, almond, myrtle –leaf milkwort, periwinkle). Acquisition rates determined by testing individual insects in quantitative PCR assays, ranging from 5.6% in N. campestris to 22.2% in P. italosignus, whereas no acquisition was recorded for L. tunetana. Successful transmissions were detected in the recipient plants exposed to P. italosignus and N. campestris, whereas no trasmissions occurred with L. tunetana. The known vector Philaenus spumarius has been included in all the experiments for validation. The systematic surveys conducted in 2016 and 2017 provided further evidence on the population dynamics and seasonal abundance of the spittlebug populations in the olive groves.

The bacterium Xylella fastidiosa (Xf) is a worldwide distributed invasive insect-borne plant pathogen, which causes lethal diseases to many economically-important crops including olives, citrus, almonds and grapes as well as numerous forest, ornamental, and uncultivated plants. The Mediterranean basin is the top supplier of olive oil with 93% of the world production and is consequently highly concerned about the recent invasion of Xf in Europe. Recently, bioeconomic models estimated putative losses induced by the spread of Xf across the European olive-producing area ranging from 1.9 to 5.2 billion euros over 50 years; however, such models did not take into account the insect vectors, which constitute a key driver of Xf spread. In the present study, we used bioclimatic species distribution models to predict the current and future climate suitability of the Mediterranean area for the main efficient or putative transmitters of Xf to olive (i.e. Philaenus spumarius, Neophilaenus campestris and Aphrophora alni). An important part of the total extent of the Mediterranean olive-producing area, mainly situated in southern Spain, Turkey and Greece, is predicted as currently poorly suitable for these vector species. Moreover, models forecast that nearly the totality of the Mediterranean olive-producing regions will likely become climatically little suitable for these vectors by 2050 due to climate change. In Europe, Xf outbreaks have occurred so far only in localities predicted as climatically suitable for these main vector species (e.g. the Apulia region of Italy) while the areas predicted as poorly suitable are still apparently Xf-free, which suggests that climate tolerances of vectors might play a main role in shaping Xf outbreaks patterns. This pattern highlights the crucial necessity of accounting for vectors when assessing risk of Xf outbreaks, and when considering vector-borne diseases in general. The risk maps presented here will have important practical application for the optimization of current and future strategies to control Xf in the Mediterranean region.

A brief review about the known-unknown-ongoing research on the mechanism of inoculation of Xylella fastidiosa by its vectors.

The phytosanitary emergency caused by the spread of Xylella fastidiosa in the Mediterranean has raised demands for a better understanding of the ecology of its presumed and candidate insect vectors. Here, we present the results of a two-year survey carried out in olive groves across southern, eastern and Central Spain and northeastern Portugal. Several sampling methods were tested and compared to select the most appropriate to estimate population levels of potential vectors of X. fastidiosa. The spittlebugs Philaenus spumarius and Neophilaenus campestris (Hemiptera: Aphrophoridae) were the main species associated with olive groves. Both species were widely present on herbaceous ground vegetation within the olive groves; P. spumarius mainly associated with Asteraceae and N. campestris with Poaceae. Due to the patchy distribution of spittlebugs within the olive groves, sweep nets were the most effective and least time-consuming sampling method for the estimation of population size both in the ground cover and tree canopies. Trends in population density showed that spittlebugs can be abundant on ground vegetation but very rare on olive canopies. Spittlebugs disperse in late spring to non-cultivated hosts that act as natural reservoirs. In late fall, adults return to the olive groves for oviposition. However, olive trees may act as transient hosts for spittlebugs and high population densities of these insect vectors should be avoided in areas where X. fastidiosa is present., This work has been financially supported by European Union Horizon 2020 research and innovation program under grant agreements no. 635646 POnTE (Pest Organisms Threatening Europe), and no. 727987 XF-ACTORS (Xylella Fastidiosa Active Containment Through a multidisciplinary-Oriented Research Strategy).