Auchenorrhyncha (Hemiptera) from the families Cicadellidae, Aphrophoridae, and Cercopidae transmit Xylella fastidiosa (Wells et al. (Xanthomonadales: Xanthomonadaceae) (Redak et al., 2004). This bacterium has previously been linked to citrus variegated chlorosis and Pierce’s disease of grapevine in the Americas (Saponari et al., 2013). Since 2013, X. fastidiosa has been reported in olive, oleander, and almond trees in Italy (Saponari et al., 2013). In fact, in the Lecce Area of Apulia (Italy), X. fastidiosa subsp. pauca cause “Olive quick decline syndrome,” affecting over 8000 km² of infected territory (Serio et al., 2024). The known vectors in Europa are Philaenus spumarius L., Neophilaenus campestris Fallén, and Philaenus italosignus Drosopolous and Remane (Saponari et al., 2014; Cavalieri et al., 2019).

Vector control is one of the most important measures for preventing disease spread. In this regard, previous research has shown that control strategies based on chemical insecticides and physical methods can reduce vector populations, particularly those of P. spumarius (Morelli et al., 2021). However, to reduce the negative impact of the synthetic insecticides, while preserving agricultural ground cover, safer products and methods should be developed.

There is currently very little knowledge on the natural enemies affecting X. fastidiosa’s vector populations in Europe (Val Nolan, 1956; Phillipson, 1960; Whittaker, 1970; Harper & Whittaker, 1976). Recently, some authors have analysed the control potential of certain predator groups coexisting in both space and time within particular agroecosystems, mainly in olive groves. For example, the predator hemipteran Zelus renardii Kolenati (Reduviidae) was recommended by Liccardo et al. (2020) as a potential candidate for controlling P. spumarius using an inundation method. Additionally, in the laboratory, Benhadi-Marín et al. (2020) investigated the differences in prospective predatory effectiveness of the spiders Araniella cucurbitina Clerck and Synaema globosum Fabricius with P. spumarius as the prey. They found differences in the functional response of the two spider species, with S. globosum exhibiting higher predation efficiency in locations with elevated prey densities.

Spiders (Araneae) are among the most important predators of other arthropods in agroecosystems, and several studies have demonstrated its key effect in the natural control of phytophagous insects (Marc et al., 1999). In addition to their high consumption rates, spiders induce other effects, such as dislodging or promoting behavioural changes, that can reduce pest populations (Michalko et al., 2019). Although predation of spiders on X. fastidiosa vectors (mainly P. spumarius) in the field has been previously reported, the data available is limited and is often based on punctual observations (Phillipson, 1960; Harper & Whittaker, 1976; Rodrigues et al., 2022).

Recently, Rodrigues et al. (2022) reported the predation of a lynx spider of the genus Oxyopes on P. spumarius, using a PCR-based diagnostic method developed to detect generalist predators. Molecular techniques and PCR-based diagnostic methods have grown in prominence in the analysis of trophic interactions in recent years (Liu et al., 2020; Uiterwaal & DeLong, 2020). Additionally, trophic interactions have been revealed using metabarcoding, a method that combines high-throughput sequencing (HTS) with DNA barcoding to identify food remnants in faeces or gut contents (Deagle et al., 2019). DNA metabarcoding of gut content has proven to be a valuable tool for characterising trophic interactions, estimating the relative importance of intraguild prey or pest species and identifying prey diets in spiders, thus revealing new insights into biological control by this group of predators (Hambäck et al., 2021; Saqib et al., 2021; Ortiz et al., 2022)

In the present study, we determined which species and specific guilds of spiders prey on vectors of X. fastidiosa in agroecosystems in southwestern Spain using a gut content metabarcoding approach.

Among the 29 spiders analysed, 26 (89.7%) produced fruitful amplification. For eight specimens (Cheiracanthium sp., Cheiracanthium erraticum Walckenaer, Runcinia grammica C. L. Koch, Zelotes tenuis L. Koch, Zelotes sp., Monaeses paradoxus Lucas, Gibbaranea bituberculata Walckenaer, Pelecopsis mengei Simon), no prey were detected.

A total of 54 ASVs and 7.9×10⁵ reads were obtained corresponding to 7 prey orders, 19 families and 30 species (Table 2). Neophilaenus campestris was detected in two spider species, namely Enoplognatha caricis Fickert and Thanatus vulgaris Simon, with 3.4 and 6.9% of the captured specimens, respectively. In addition, L. coleoptrata was preyed on by Haplodrassus macellinus Thorell, with 3.4% of the specimens. Philaenus spumarius were not detected in the samples. As for the most frequent prey, 43.3% of the total prey detected were spider species, with 23.3% hemipterans (6.7% of which were aphrophorids), and 13.3% dipterans (including two parasitoid species; Table 2). Enoplognatha species are known for their mobility, frequently move within inflorescences and can drop on a silken thread and move out of the plant either to the ground or to another plant (Greco & Kevan, 1999), increasing their chances of encountering the vector nymphs, a feature rarely found in other web-building spiders. The other two spiders preying on aphrophorids (T. vulgaris and H. macellinus included in the “other-hunters” and “ground-hunters” guilds, respectively) do not build webs. Instead, they actively move across the foliage and ground in search of prey (Uetz, 1992). These hunting strategies are more effective for less mobile prey, such as vector nymphs. Limited data have been previously reported regarding natural predators on European X. fastidiosa vectors. Punctual observations of birds (Val Nolan, 1956; Whitaker, 1970), carabids, phalangids and spiders (Harper & Whitaker, 1976; Rodrigues et al., 2022) preying on aphrophorids (mainly P. spumarius) have been previously reported. Among spiders, the Araneidae, Lycosidae and Oxyopidae families have been previously identified as predators of P. spumarius (Harper & Whitaker, 1976; Rodrigues et al., 2022). This is the first report of predation by spiders from Philodromidae and Gnaphosidae, expanding the range of spider species and families capable of exerting control of X. fastidiosa vectors.

In this study, N. campestris was only detected in vineyards, and L. coleoptrata was detected only in citrus orchards. No aphrophorids were detected in olive orchards. “Other hunters”, “ground hunters” and “space-web” spiders preyed on aphrophorids while vectors were not detected in the guts of “ambush hunters” or “orb-web” guilds (Figure 1). The number of taxa detected in the gut content per spider varied significantly among crops (LRT χ2=17.9; p<0.001) and spider guilds (LRT χ2=25.9; p<0.001). In this context, the mean number of prey taxa per spider was significantly higher in vineyards (4.2±2.7) and citrus orchards (2.9±0.7) than in olive orchards (1.2±0.3; z=4.03 and −2.8; p<0.05). Among spider guilds, the mean number of prey taxa was lowest for “ambush hunters” (0.4±0.2) with significant differences with respect to highest obtained for “other hunters” group (3.5±1.2; z=2.9; p<0.02). Finally, the other three guilds presented no significant differences with mean number of prey taxa ranging from 1.0±0.6 and 1.0±0.7 for “orb-web” and “ground hunter” spiders, respectively, to 3.0±1.5 for “sheet-web” spiders.

Rosas-Ramos et al. (2018) documented the predominance of Philodromidae spiders in Spanish vineyards. Our results suggest that this spider family (represented by T. vulgaris) is involved in the natural control of aphrophorids in vineyards, consistent with their observed abundance. Previous research has shown that Philodromidae, Salticidae and Araneae are among the most abundant spider families in olive orchards in the south of the Iberian Peninsula (Morris et al., 1999; Cárdenas et al., 2012). In addition, Rodrigues et al. (2022) detected P. spumarius in the gut of linx spider Oxyopes sp. in a survey carried out in olive orchards in northeastern Portugal. Our findings align with these studies, reinforcing the role of these spider families as natural predators of aphrophorids. However, our study did not detect aphrophorids in the gut content of spiders in the olive orchards of southwest Spain. This absence may be due to regional differences in prey availability or spider behaviour. To better understand these dynamics, more representative and intensive sampling-based studies are required.