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.

Probing behavior of the corn leafhopper Dalbulus maidis on susceptible and resistant maize hybrids

The corn leafhopper Dalbulus maidis is the main vector of the pathogens that cause corn stunt, a major disease of maize in the Americas. As host resistance is an efficient tool to control diseases, the findings of a previous report showed that some corn hybrids are resistant to D . maidis . In this work, we assessed the probing behavior of D . maidis on susceptible and resistant corn hybrids using EPG (Electrical Penetration Graph) technology. Fifteen-day-old females were monitored for 20 hours, with access to hybrids DK390, DK670, DK79-10, and DK72-10. Hybrids DK390 and DK72-10 showed resistance to D . maidis in phloem, since insects feeding on these hybrids presented more salivation events in phloem without subsequent ingestion, which are seen as failed attempts to ingest. A reduction of the total duration of phloem ingestion was observed, and accordingly of the time spent by insects with access to these hybrids on xylem ingestion. The hybrid DK390 also had mesophyll resistance, seen as less probing time and a higher number of probes of short duration. These findings support and are consistent with previous research, providing useful information to characterize maize hybrids resistant to D . maidis , and so to corn stunt.

Effect of Sugarcane Cultivars Infected with Sugarcane Yellow Leaf Virus (ScYLV) on Feeding Behavior and Biological Performance of Melanaphis sacchari (Hemiptera: Aphididae)

Sugarcane yellow leaf virus (ScYLV), Polerovirus, Luteoviridae, is one of the main viruses that infect sugarcane worldwide. The virus is transmitted by the aphid Melanaphis sacchari in a persistent, circulative manner. To better understand the interactions between ScYLV, sugarcane genotypes and M. sacchari, we explored the effect of sugarcane cultivars on the feeding behavior and biological performance of the vector. The number of nymphs, adults, winged, total number of aphids and dead aphids was assayed, and an electrical penetration graph (EPG) was used to monitor the stylet activities. Multivariate analysis showed changes in the vector’s behavior and biology on cultivars, identifying specific groups of resistance. In the cultivar 7569, only 5.5% of the insects were able to stay longer on sustained phloem ingestion, while in the other seven cultivars these values varied from 20% to 60%. M. sacchari showed low phloem activities in cultivars 7569 and Bio266. Overall, cultivar 7569 showed the worst biological performance of aphids, with the insects presenting mechanical difficulties for feeding and a shorter duration of the phloem period, and thus being considered the most resistant. We conclude that ScYLV virus infection in different sugarcane cultivars induced specific changes in the host plant, modifying the behavior of its main vector, which may favor or impair virus transmission.

RNA-Seq and Electrical Penetration Graph Revealed the Role of Grh1-Mediated Activation of Defense Mechanisms towards Green Rice Leafhopper (Nephotettix cincticeps Uhler) Resistance in Rice (Oryza sativa L.)

The green rice leafhopper (GRH, Nephotettix cincticeps Uhler) is one of the most important insect pests causing serious damage to rice production and yield loss in East Asia. Prior to performing RNA-Seq analysis, we conducted an electrical penetration graph (EPG) test to investigate the feeding behavior of GRH on Ilpum (recurrent parent, GRH-susceptible cultivar), a near-isogenic line (NIL carrying Grh1) compared to the Grh1 donor parent (Shingwang). Then, we conducted a transcriptome-wide analysis of GRH-responsive genes in Ilpum and NIL, which was followed by the validation of RNA-Seq data by qPCR. On the one hand, EPG results showed differential feeding behaviors of GRH between Ilpum and NIL. The phloem-like feeding pattern was detected in Ilpum, whereas the EPG test indicated a xylem-like feeding habit of GRH on NIL. In addition, we observed a high death rate of GRH on NIL (92%) compared to Ilpum (28%) 72 h post infestation, attributed to GRH failure to suck the phloem sap of NIL. On the other hand, RNA-Seq data revealed that Ilpum and NIL GRH-treated plants generated 1,766,347 and 3,676,765 counts per million mapped (CPM) reads, respectively. The alignment of reads indicated that more than 75% of reads were mapped to the reference genome, and 8859 genes and 15,815,400 transcripts were obtained. Of this number, 3424 differentially expressed genes (DEGs, 1605 upregulated in Ilpum and downregulated in NIL; 1819 genes upregulated in NIL and downregulated in Ilpum) were identified. According to the quantile normalization of the fragments per kilobase of transcript per million mapped reads (FPKM) values, followed by the Student’s t-test (p < 0.05), we identified 3283 DEGs in Ilpum (1935 upregulated and 1348 downregulated) and 2599 DEGs in NIL (1621 upregulated and 978 downregulated) with at least a log2 (logarithm base 2) twofold change (Log2FC ≥2) in the expression level upon GRH infestation. Upregulated genes in NIL exceeded by 13.3% those recorded in Ilpum. The majority of genes associated with the metabolism of carbohydrates, amino acids, lipids, nucleotides, the activity of coenzymes, the action of phytohormones, protein modification, homeostasis, the transport of solutes, and the uptake of nutrients, among others, were abundantly upregulated in NIL (carrying Grh1). However, a high number of upregulated genes involved in photosynthesis, cellular respiration, secondary metabolism, redox homeostasis, protein biosynthesis, protein translocation, and external stimuli response related genes were found in Ilpum. Therefore, all data suggest that Grh1-mediated resistance against GRH in rice would involve a transcriptome-wide reprogramming, resulting in the activation of bZIP, MYB, NAC, bHLH, WRKY, and GRAS transcription factors, coupled with the induction of the pathogen-pattern triggered immunity (PTI), systemic acquired resistance (SAR), symbiotic signaling pathway, and the activation of genes associated with the response mechanisms against viruses. This comprehensive transcriptome profile of GRH-responsive genes gives new insights into the molecular response mechanisms underlying GRH (insect pest)–rice (plant) interaction.

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.

Trehalose and glucose levels regulate feeding behavior of the phloem-feeding insect, the pea aphid Acyrthosiphon pisum Harris

Trehalose serves multifarious roles in growth and development of insects. In this study, we demonstrated that the high trehalose diet increased the glucose content, and high glucose diet increased the glucose content but decreased the trehalose content of Acyrthosiphon pisum. RNA interference (RNAi) of trehalose-6-phosphate synthase gene (ApTPS) decreased while RNAi of trehalase gene (ApTRE) increased the trehalose and glucose contents. In the electrical penetration graph experiment, RNAi of ApTPS increased the percentage of E2 waveform and decreased the percentage of F and G waveforms. The high trehalose and glucose diets increased the percentage of E2 waveform of A. pisum red biotype. The correlation between feeding behavior and sugar contents indicated that the percentage of E1 and E2 waveforms were increased but np, C, F and G waveforms were decreased in low trehalose and glucose contents. The percentage of np, E1 and E2 waveforms were reduced but C, F and G waveforms were elevated in high trehalose and glucose contents. The results suggest that the A. pisum with high trehalose and glucose contents spent less feeding time during non-probing phase and phloem feeding phase, but had an increased feeding time during probing phase, stylet work phase and xylem feeding phase.

Antixenosis in Glycine max (L.) Merr against Acyrthosiphon pisum (Harris)

To reveal the antixenosis potential against the pea aphid Acyrthosiphon pisum (Harris) (Hemiptera: Aphididae) we analyzed the pea aphid survival and probing behavior, and the quantitative and qualitative variation of flavonoids in the leaves of selected soybean Glycine max (L.) Merr (Fabaceae) cultivars ‘Aldana’, ‘Annushka’, ‘Augusta’, ‘Madlen’, ‘Mavka’, ‘Simona’, ‘Violetta’, and ‘Viorica’. Aphid survival was drastically impeded on all cultivars. The electronic monitoring of aphid probing using the Electrical Penetration Graph (EPG) technique revealed that on all soybean cultivars, A. pisum readily probed into leaf tissues but the probes were usually terminated before reaching vascular tissues, which demonstrates the activity of antixenosis mechanisms in peripheral tissues epidermis and/or mesophyll in soybean leaves. The potency of antixenosis factors differed among soybean cultivars, which was reflected in differences in aphid survival and frequency and duration of phloem sap ingestion. Seven flavonoids were found: apigenin, daidzein, genistein, glycitein, isorhamnetin, kaempferol, and rutin, which occurred in different amount and proportion in individual cultivars. The content of apigenin and genistein in all soybean cultivars studied probably made them relatively unacceptable to A. pisum. Kaempferol in ‘Aldana’ might be responsible for the observed strong antixenosis resistance of this cultivar to A. pisum. The results of our survey provide the first detailed data that can be used for future studies.

The Source of Rag5-Mediated Resistance to Soybean Aphids Is Located in the Stem

The soybean aphid (Aphis glycines) continues to threaten soybean production in the United States. A suite of management strategies, such as planting aphid-resistant cultivars, has been successful in controlling soybean aphids. Several Rag genes (resistance against A. glycines) have been identified, and two are currently being deployed in commercial soybean cultivars. However, the mechanisms underlying Rag-mediated resistance are yet to be identified. In this study, we sought to determine the nature of resistance conferred by the Rag5 gene using behavioral, molecular biology, physiological, and biochemical approaches. We confirmed previous findings that plants carrying the Rag5 gene were resistant to soybean aphids in whole plant assays, and this resistance was absent in detached leaf assays. Analysis of aphid feeding behaviors using the electrical penetration graph technique on whole plants and detached leaves did not reveal differences between the Rag5 plants and Williams 82, a susceptible cultivar. In reciprocal grafting experiments, aphid populations were lower in the Rag5/rag5 (Scion/Root stock) chimera, suggesting that Rag5-mediated resistance is derived from the shoots. Further evidence for the role of stems comes from poor aphid performance in detached stem plus leaf assays. Gene expression analysis revealed that biosynthesis of the isoflavone kaempferol is upregulated in both leaves and stems in resistant Rag5 plants. Moreover, supplementing with kaempferol restored resistance in detached stems of plants carrying Rag5. This study demonstrates for the first time that Rag5-mediated resistance against soybean aphids is likely derived from stems.

Un-silencing an Experimental Technique: Listening to the Electrical Penetration Graph

In scientific work, sonification (i.e., the use of nonspeech audio to convey information) is primarily thought of as a novel way to communicate post hoc research findings to lay audiences but only rarely, if ever, as a component of the research itself. This article argues that, rather than any inherent epistemological limitations of sound as a medium of scientific reasoning, this framing reflects a sociohistorical tendency to “silence” experimental techniques as they become widely adopted—both in terms of the literal silencing of noisy instrumentation and the elision of the role of sound (along with other nonvisual sensory media) in received discovery accounts. This tendency is well-illustrated by the history of electrophysiology research, particularly the case of the electrical penetration graph (EPG), an instrument used by entomologists to study how insects feed on plants. Combining participant observation, interviews, and analysis of EPG technical literature, this article recovers the unacknowledged role of sound in EPG’s exploratory stages and gradual emergence as a standard laboratory technique as well as its gradual silencing as a consequence of this standardizing process.

Impact of Temperature on Survival Rate, Fecundity, and Feeding Behavior of Two Aphids, Aphis gossypii and Acyrthosiphon gossypii, When Reared on Cotton

Aphid performance is sensitive to temperature changes. Previous studies found that Acyrthosiphon gossypii (Mordviiko) was more sensitive to high temperature than Aphis gossypii (Glover). However, the effects of high temperatures on the survival, fecundity, and feeding behavior of these two aphid adults are not clear. This study examined the effect of different temperatures (29 °C, 32 °C, and 35 °C) on the adult survival rate, fecundity, and feeding behavior of these two aphid species. Our results showed that the adverse effects of high temperatures (32 °C and 35 °C) on aphid adult survival and fecundity were greater for Ac. gossypii than Ap. gossypii. The electrical penetration graph (EPG) data showed that Ac. gossypii spent more time feeding on xylem than phloem under all temperature treatments, which contrasted with Ap. gossypii. The time of phloem ingestion by Ap. gossypii at 32 °C was significantly higher than at 29 °C, while for Ac. gossypii, this value significantly decreased when temperature increased. These feeding patterns indicate that Ac. gossypii obtains less nutrition from phloem in support of its development and fecundity. Data generated in this study will serve as the basis for predicting the effects of increased temperature on these two cotton aphids.