Scaphoideus titanus Ball feeding behaviour on three grapevine cultivars with different susceptibilities to Flavescence dorée

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 probing 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 phases. 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 preference for the Barbera variety, that appears a more suitable hosts 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 variety to FD. The mechanisms for the non-preference for Brachetto and Moscato are discussed, and a possible 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.

Effect of Soy Leaf Flavonoids on Pea Aphid Probing Behavior

Flavonoids detected in soybean Glycine max (L.) Merr. (Fabaceae) cause various alterations in the metabolism, behavior, and development of insect herbivores. The pea aphid Acyrthosiphon pisum (Harris) (Hemiptera: Aphididae) poses potential threat to soybeans, but the effect of individual flavonoids on its feeding-associated behavior is relatively unknown. We monitored probing behavior (stylet penetration activities) of A. pisum on its preferred host plant, Pisum sativum L. untreated (control) and treated with 0.1% ethanolic solutions of flavonoids apigenin, daidzein, genistein, and kaempferol. We applied the electrical penetration graph (electropenetrography, EPG) technique, which visualizes the movements of aphid stylets within plant tissues. None of the applied flavonoids affected the propensity to probe the plants by A. pisum. However, apigenin enhanced the duration of probes in non-phloem tissues, which caused an increase in the frequency and duration of stylet mechanics derailment and xylem sap ingestion but limited the ingestion of phloem sap. Daidzein caused a delay in reaching phloem vessels and limited sap ingestion. Kaempferol caused a reduction in the frequency and duration of the phloem phase. Genistein did not affect aphid probing behavior. Our findings provide information for selective breeding programs of resistant plant cultivars to A. pisum.

Synthesis of novel phytol-derived γ-butyrolactones and evaluation of their biological activity

The synthesis of phytol-derived γ-butyrolactones as well as their evaluation for deterrent activity towards peach-potato aphid Myzus persicae and antiproliferative activity against four selected cancer cell lines are reported. Products were obtained in good yields (19–96%) and their structures were fully characterized by spectroscopic data (NMR, HRMS). Four synthesized δ-halo-γ-lactones (4–7) are new and have not been previously described in the literature. In the choice test phytol (1) appeared deterrent to M. persicae, whereas modifications of its structure did not cause the avoidance of the treated leaves by the aphids. In contrast, aphids were attracted to the leaves treated with the new trans-δ-chloro-γ-lactone (6). Electrical Penetration Graph (EPG) technique applied to explore the aphid probing and feeding activity revealed that neither phytol nor lactone 6 affected aphid probing and the consumption of phloem sap, which means that both phytol and the lactone 6 might have acted as postingestive modifiers of aphid behavior. The results of in vitro antitumor assays showed that obtained phytol derivatives exhibit cytotoxic activity against studied cancer cell lines (leukemia, lung and colon carcinoma and its doxorubicin resistant subline). Halolactones 4–6 were identified as the compounds, which arrest cell cycle of leukemia cells mainly in G2/M and S phases.

Semipersistently Transmitted, Phloem Limited Plant Viruses Are Inoculated during the First Subphase of Intracellular Stylet Penetrations in Phloem Cells

The green peach aphid Myzus persicae Sulzer is the main vector of the semipersistently transmitted and phloem-limited Beet yellows virus (BYV, Closterovirus). Studies monitoring the M. persicae probing behavior by using the Electrical penetration graphs (EPG) technique revealed that inoculation of BYV occurs during unique brief intracellular punctures (phloem-pds) produced in companion and/or sieve element cells. Intracellular stylet punctures (or pds) are subdivided in three subphases (II-1, II-2 and II-3), which have been related to the delivery or uptake of non-phloem limited viruses transmitted in a non-persistent or semipersistent manner. As opposed to non-phloem limited viruses, the specific pd subphase(s) involved in the successful delivery of phloem limited viruses by aphids remain unknown. Therefore, we monitored the feeding process of BYV-carrying M. persicae individuals in sugar beet plants by the EPG technique and the feeding process was artificially terminated at each phloem-pd subphase. Results revealed that aphids that only performed the subphase II-1 of the phloem-pd transmitted BYV at similar efficiency than those allowed to perform subphase II-2 or the complete phloem-pd. This result suggests that BYV inoculation occurs during the first subphase of the phloem-pd. The specific transmission mechanisms involved in BYV delivery in phloem cells are discussed.

Repellent and Antifeedant Activities of Citral-Derived Lactones against the Peach Potato Aphid

Citral is well known for its antimicrobial, antifungal, and insecticidal activities. Natural sesquiterpene α-methylenelactones also exhibit a broad spectrum of biological activities. The aim of the study was to explore the effect of structural changes to citral molecules on citral behavior-modifying activity towards Myzus persicae. Specifically, the effects of the introduction of a γ-lactone moiety and methylene groups in α and γ positions of the lactone ring were investigated. The lactones were obtained in five-step (saturated lactone and γ-methylenelactone) or six-step (α-methylenelactone and α,γ-dimethylenelactone) syntheses from citral. The synthetic procedures and physical and spectral data of the lactones are presented. The settling behavior of freely moving aphids in choice and no-choice situations was monitored. The probing behavior of tethered M. persicae using the Electrical Penetration Graph (EPG) technique was also analyzed. Citral appeared a strong repellent and pre-ingestive and ingestive probing deterrent to M. persicae. The incorporation of a lactone moiety caused the loss of the repellent activity. α-Methylenelactone inhibited aphid settling and probing activities at pre-ingestive and ingestive phases. The saturated γ-lactone and α,γ-dimethylenelactone were the settling post-ingestive deterrents to M. persicae, which did not affect aphid probing activity. γ-Methylenelactone did not affect aphid behavior.

Effect of Naringenin and Its Derivatives on the Probing Behavior of Myzus persicae (Sulz.)

Substances that alter insect behavior have attracted a lot of attention as potential crop protection agents. Naringenin (5,7,4′-trihydroxyflavanone) is a naturally occurring bioactive flavanone. We evaluated the influence of naringenin on aphid activities during individual phases of probing and feeding and the effect of structural modifications of naringenin on its activity towards aphids. We monitored the probing behavior of Myzus persicae (Sulz.) (Hemiptera: Aphididae) using the Electrical Penetration Graph (EPG) technique. The chemical modifications were the substitution of hydrogen atoms with methyl, ethyl or pentyl groups and the replacement of the carbonyl group in naringenin and its derivatives with an oxime moiety. Depending on the substituents, the activity of naringenin-derived compounds varied in potency and mode of action. Naringenin was an attractant of moderate activity, which enhanced sap ingestion. The naringenin derivative with two methyl groups—7,4′-di-O-methylnaringenin—was a deterrent, which hindered aphid probing in non-phloem tissues. Naringenin oxime derivatives with methyl substituents—7,4′-di-O-methylnaringenin oxime, 7-O-methylnaringenin oxime, and 5,7,4′-tri-O-methylnaringenin oxime—and the derivative with a pentyl substituent—7-O-pentylnaringenin oxime—were strong attractants which stimulated aphid probing in non-phloem tissues and the ingestion of phloem sap.

Comparative Analysis of Stylet Penetration Behaviors of Eriosoma lanigerum (Hemiptera: Aphididae) on Main Apple Cultivars in China

The preference and behavioral mechanism of the feeding behavior of the woolly apple aphid, Eriosoma lanigerum (Hausmann), on Chinese apple cultivars was investigated using the electrical penetration graph (EPG) technique. Feeding of adult aphids was assessed for the four main cultivars of apple in China (Red Fuji, Ralls Genet, Starkrimson, and Jonagold) over an 8-h monitoring period, what that of fourth-instar nymphs was assessed for 8 h on Jonagold only. EPG waveforms representing the details of probing behaviors, parameters were measured for analysis. We found that at the nonphloem phase, the total duration of the nonpenetration (np) waveform and the total duration of the stylet pathway phase (the ‘C wave’) on Ralls Genet and Starkrimson were significantly longer than on Red Fuji. In addition, the phloem-feeding phase, the total duration of ‘release of saliva into the phloem’, (the E1 waveform) was significantly longer on Ralls Genet and Jonagold than on Red Fuji. The total duration of the ‘ingestion from sieve cells’ phase (the E2 waveform) on Red Fuji, meanwhile, was significantly longer than on Ralls Genet or Starkrimson. At the same time, when combined with EPG parameters, Red Fuji was found to have no obvious resistance to woolly apple aphid and to be a susceptible cultivar. This experiment also recorded the feeding waveform of fourth-instar aphid nymphs. Based on the phloem and nonphloem parameters recorded in this study, it was inferred that adult aphids are more likely than nymphs to feed on apple trees.

New insight into the behaviour modifying activity of two natural sesquiterpenoids farnesol and nerolidol towards Myzus persicae (Sulzer) (Homoptera: Aphididae)

The effect of structurally related sesquiterpenoids (E,E)-farnesol and cis-nerolidol on the host-plant selection behaviour of the peach potato aphid Myzus persicae (Sulz.) was evaluated using electrical penetration graph (EPG) technique. No repellent effects of (E,E)-farnesol and (Z)-nerolidol to M. persicae were found but aphid probing activities on (E,E)-farnesol- and cis-nerolidol-treated plants were restrained. During non-phloem phases of probing, neither (E,E)-farnesol nor (Z)-nerolidol affected the cell puncture activity. On (E,E)-farnesol-treated plants, the total duration of phloem phase, the mean duration of individual sustained ingestion periods were significantly lower, and the proportion of phloem salivation was higher than on control plants. On (Z)-nerolidol-treated plants, the occurrence of the first phloem phase was delayed, and the frequency of the phloem phase was lower than on control plants. The freely moving aphids were reluctant to remain on (E,E)-farnesol- and (Z)-nerolidol-treated leaves for at least 24 h after exposure. (E,E)-Farnesol and (Z)-nerolidol show complementary deterrent properties, (E,E)-farnesol showing ingestive and post-ingestive activities and nerolidol showing pre-ingestive, ingestive, and post-ingestive deterrent activities.

Aphid-Triggered Changes in Oxidative Damage Markers of Nucleic Acids, Proteins, and Lipids in Maize (Zea mays L.) Seedlings

Prior experiments illustrated reactive oxygen species (ROS) overproduction in maize plants infested with bird-cherry-oat (Rhopalosiphum padi L.) aphids. However, there is no available data unveiling the impact of aphids feeding on oxidative damages of crucial macromolecules in maize tissues. Therefore, the purpose of the current study was to evaluate the scale of oxidative damages of genomic DNA, total RNA and mRNA, proteins, and lipids in seedling leaves of two maize genotypes (Złota Karłowa and Waza cvs—susceptible and relatively resistant to the aphids, respectively). The content of oxidized guanosine residues (8-hydroxy-2′-deoxyguanosine; 8-OHdG) in genomic DNA, 8-hydroxyguanosine (8-OHG) in RNA molecules, protein carbonyl groups, total thiols (T-SH), protein-bound thiols (PB-SH), non-protein thiols (NP-SH), malondialdehyde (MDA) and electrolyte leakage (EL) levels in maze plants were determined. In addition, the electrical penetration graphs (EPG) technique was used to monitor and the aphid stylet positioning and feeding modes in the hosts. Maize seedlings were infested with 0 (control), 30 or 60 R. padi adult apterae per plant. Substantial increases in the levels of RNA, protein and lipid oxidation markers in response to aphid herbivory, but no significant oxidative damages of genomic DNA, were found. Alterations in the studied parameters were dependent on maize genotype, insect abundance and infestation time.

Potential use of electrical penetration graph (EPG) technology for biosecurity incursion response decision making

Sap-sucking insects pose a significant biosecurity risk as they can cause direct damage to plants and may also vector serious plant pathogens. However, there is a paucity of tools for rapidly estimating the host range of such insects during a biosecurity incursion. The Electrical Penetration Graph (EPG) technique has been developed over the past 50 years for analysing, quantifying, and comparing the feeding behaviours of both phloem and xylem sap-sucking insects. Electrically generated waveforms associated with insects’ probing and feeding behaviours may allow a rapid assessment of the potential host range of invading sap sucking insects. This paper reviews the development and use of EPG, assesses potential for host-range testing of recent incursions, analyses when and how the EPG tool could be applied for response, and identifies seven points where EPG could aid biosecurity incursion response decision making. Three recent biosecurity responses could have benefited from EPG.