Electropenetrography Study of Euschistus heros (F.) (Heteroptera: Pentatomidae) Nymphs Feeding on Vegetative Soybean Structures.

The study aimed to understand the feeding behavior of Neotropical brown stink bug nymphs Euschistus heros (F.) on soybean plants during vegetative stage through electropenetrography (EPG) technique. Three distinct phases were identified: non-feeding, pathway, and ingestion. Waveforms representing these phases were consistent across nymphal instars and plant structures, and named Np, Eh1, and Eh2, respectively. Biological interpretations of the waveforms were proposed by integrating visual observations, comparisons with adult waveforms, and histological studies. The waveforms Np, Eh1, and Eh2 were associated with resting/walking, initial stylet-plant contact, and xylem sap ingestion, respectively. Notably, nymphs showed a higher number of Eh1 events and longer durations when feeding on petioles compared to leaves, particularly in younger instars. However, differences between instars diminished in older nymphs. Fifth instars consistently exhibited the highest Eh1 values, and displayed longer xylem ingestion durations compared to other instars. Second instars demonstrated increased xylem ingestion events on petioles compared to leaves. Across plant structures, on petioles, nymphs generally showed longer xylem ingestion durations than on leaf surfaces. Fifth instar consistently had the longest ingestion durations overall. Additionally, statistical differences in xylem ingestion duration were observed between instars within each plant structure, with fifth instars displaying the longest durations. These findings offer valuable insights into the feeding behavior of E. heros nymphs, which could inform the development of more effective pest management strategies for soybean crops.

Bt Soybean Cry1Ac Does Not Affect Development, Reproduction, or Feeding Behavior of Red-Banded Stink Bug Piezodorus guildinii (Hemiptera: Pentatomidae).

Bt soybean cultivation is increasing worldwide. The Cry1Ac protein expressed in Bt soybean efficiently controls several lepidopteran pests. The stink bug, Piezodorus guildinii (Westwood), a major pest for soybean in the Americas, is not controlled by Bt crops, although possible sub-lethal effects may occur. Even if there were no negative effects for sting bug, ingesting toxins could affect its bio-controllers. We tested through ELISA detection if P. guildinii ingests Cry1Ac from Bt soybean and possible effects on its development, reproduction, survival, and feeding behavior. Biological traits were evaluated under controlled conditions of nymphs and adults feeding on pods of near-isogenic cultivars DM5958iPRO (Bt) and DM59i (non-Bt). Feeding behavior was recorded using an AC-DC electropenetrography (EPG) device. Results indicated that P. guildinii ingested the Cry1Ac protein; however, nymphal period and accumulated survival percentage did not differ between cultivars. Feeding on Bt soybean pods did not affect fecundity (i.e., number of egg masses and eggs/female) nor egg viability. Different feeding behaviors were only detected on the pathway phase (stylet penetration into plant tissue), which was more pronounced in the Bt cultivar. However, the total duration of the feeding activities on seeds was numerically higher (ca. 2X) on Bt plants compared to non-Bt. This is the first study to demonstrate that P. guildinii does ingest the Cry1Ac protein and excrete it without being absorbed, probably explaining the lack of direct adverse effects on its biological parameters. EPG could indicate that Bt soybean plants might be less palatable than non-Bt to red-banded stink bug.

Electropenetrography study of Euschistus heros (F.) (Heteroptera: Pentatomidae) nymphs feeding at different sites of the soybean pod.

This study aimed to characterize and correlate the stylet penetration behaviors of nymphs of the Neotropical brown stink bug, Euschistus heros (F.), on immature soybean pods. Waveforms were obtained using electropenetrography (EPG). The findings revealed that the nymphs exploited the xylem vessels and the seed tegument or endosperm. Primarily 4 phases were characterized: nonfeeding, pathway, salivation, and ingestion. The waveforms of each phase were similar in appearance across instars. The biological meanings of waveforms were based on visual observations, comparison with waveforms of adults, and histological studies. Np represents the insect resting or walking on soybean pod surface. Eh1 represents the first contact between the mouthparts (stylets) and plant tissue. Eh2 represents xylem sap ingestion, and Eh3 represents seed activities (including tegument and endosperm). The number of waveform events did not differ among instars for all waveforms. However, for Eh3, fifth instars performed more activities than other instars. The second instars had the smallest value, and third and fourth instars had intermediate values. For total duration, all waveforms differed among instars. Np duration was shorter for third compared with second and fourth instars and intermediate for fifth instar. For Eh1, second and third instars had the longest duration (1.5× to 2× greater) compared with fourth and fifth instars. For Eh2 and Eh3, the second-instar showed the longest (~2× greater) and shortest durations, respectively. Overall, this study provides important insights into the feeding behavior of E. heros nymphs so that effective pest management programs can be developed to contain this pest.

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.

Evaluating resistance of the soybean block technology cultivars to the Neotropical brown stink bug, Euschistus heros (F.).

Laboratory studies were conducted with the Neotropical brown stink bug, Euschistus heros (F.), to evaluate nymphal and adult biology on immature pods of soybean, Glycine max (L.) Merrill (Fabaceae), bearing the block technology (resistant to stink bug damage - cvs. BRS 391, BRS 543 RR and BRS 1003 IPRO) compared to a susceptible cultivar (BRS 5601 RR). Results indicated that nymphs' developmental time and survivorship were similar on all cultivars tested. The same was observed for adult survivorship and reproductive performance. However, data from electropenetrography (EPG) demonstrated that adults of E. heros spent significantly less time in feeding activities on resistant plants compared to the susceptible one. Large differences were observed in feeding activities on seeds; on resistant plants, the insects dedicated a shorter period of time to feed on seed endosperm than on BRS 5601. In addition, when bugs fed on seeds of block cultivars, the majority of probes were composed of only laceration/maceration activities (Eh3a waveform) without ingestion events of the cell contents (Eh3b waveform). In contrast, on the susceptible cultivar, Eh3a waveform events were repeated much more frequently (3-5X) with more probes also containing ingestion of seed contents. These results suggest that the soybean cultivars bearing the block technology presented a lower preference (antixenosis) by the bugs with fewer feeding activities, primarily in the seed endosperm, compared to the susceptible one tested.

Plant Resistance in Some Modern Soybean Varieties May Favor Population Growth and Modify the Stylet Penetration of Bemisia tabaci (Hemiptera: Aleyrodidae).

Complaints of severe damage by whiteflies in soybean fields containing genetically engineered (GE) varieties led us to investigate the role of transgenic soybean varieties expressing resistance to some insects (Cry1Ac Bt toxin) and to herbicide (glyphosate) on the population growth and feeding behavior of Bemisia tabaci (Gennadius) MEAM1 (Hemiptera: Aleyrodidae). In the laboratory, the whiteflies reared on the GE Bt soybeans had a net reproductive rate (R0) 100% higher and intrinsic rate of population increase (rm) 15% higher than those reared on non-GE soybeans. The increased demographic performance was associated with a higher lifetime fecundity. In electrical penetration graphs, the whiteflies reared on the GE soybeans had fewer probes and spent 50% less time before reaching the phloem phase from the beginning of the first successful probe, indicating a higher risk of transmission of whitefly-borne viruses. Data from Neotropical fields showed a higher population density of B. tabaci on two soybean varieties expressing glyphosate resistance and Cry1Ac Bt toxin. These results indicate that some GE soybean varieties expressing insect and herbicide resistances can be more susceptible to whiteflies than non-GE ones or those only expressing herbicide resistance. Most likely, these differences are related to varietal features that increase host-plant susceptibility to whiteflies. Appropriate pest management may be needed to deal with whiteflies in soybean fields, especially in warm regions, and breeders may want to consider the issue when developing new soybean varieties.

Unveiling the Feeding Behavior of Tibraca limbativentris (Hemiptera: Pentatomidae) on Rice Using an Electropenetrography Waveform Library.

The rice stalk stink bug, Tibraca limbativentris Stål, damages plant stalks while feeding, making it one of the most important rice pests in South America. Because the feeding behavior of T. limbativentris has not yet been studied in rice, we investigated T. limbativentris stylet penetration (probing) in rice stalks. A waveform library was created using the new AC-DC EPG monitor with different levels of input resistance (Ri). Six different waveforms were recorded and correlated via histological studies and grouped into three phases: non-probing waveforms (Z and Np), pathway waveforms (Tl1), and ingestion waveforms (Tl2 and Tl3). The Z waveform was observed when the stink bug was standing still on the plant surface, Np when the stink bug was walking on plant surface, Tl1 was associated with stylet insertion and deep penetration into the plant tissue, and Tl2 when the stink bug was feeding on xylem vessels. The Tl3 waveform was associated with the rupture of stalk cells and was divided into two subtypes (Tl3a and Tl3b). The Tl3a waveform probably represents cell laceration with combined enzymatic maceration of stalk tissues, while Tl3b represents a short ingestion period of macerated tissues. Tibraca limbativentris uses two strategies to feed on rice stalks: a salivary sheath for feeding on xylem vessels and cell rupture (laceration and maceration) for feeding on parenchyma cells. Our study provides crucial benchmark definitions of waveforms. Future studies can now compare effects of treatments on stink bug feeding, to ultimately improve management of this pest in rice.

Foliar Spraying of Tomato Plants with Systemic Insecticides: Effects on Feeding Behavior, Mortality and Oviposition of Bemisia tabaci (Hemiptera: Aleyrodidae) and Inoculation Efficiency of Tomato Chlorosis Virus.

Tomato chlorosis virus (ToCV) is a phloem-limited crinivirus transmitted by whiteflies and seriously affects tomato crops worldwide. As with most vector-borne viral diseases, no cure is available, and the virus is managed primarily by the control of the vector. This study determined the effects of the foliar spraying with the insecticides, acetamiprid, flupyradifurone and cyantraniliprole, on the feeding behavior, mortality, oviposition and transmission efficiency of ToCV by B. tabaci MEAM1 in tomato plants. To evaluate mortality, oviposition and ToCV transmission in greenhouse conditions, viruliferous whiteflies were released on insecticide-treated plants at different time points (3, 24 and 72 h; 7 and 14 days) after spraying. Insect mortality was higher on plants treated with insecticides; however, only cyantraniliprole and flupyradifurone differed from them in all time points. The electrical penetration graph (DC-EPG) technique was used to monitor stylet activities of viruliferous B. tabaci in tomato plants 72 h after insecticide application. Only flupyradifurone affected the stylet activities of B. tabaci, reducing the number and duration of intracellular punctures (pd) and ingestion of phloem sap (E2), a behavior that possibly resulted in the lower percentage of ToCV transmission in this treatment (0-60%) in relation to the control treatment (60-90%) over the periods evaluated. Our results indicate that flupyradifurone may contribute to management of this pest and ToCV in tomato crops.

Electropenetrographic Comparison of Feeding Behavior of Dichelops furcatus (Hemiptera: Heteroptera: Pentatomidae) on Soybean and Spring Cereals.

We used electropenetrography to quantify and compare counts and durations of selected waveforms, produced by adult females of the stink bug Dichelops furcatus (F.). Insects fed on immature soybean pods and immature seed heads of four spring cereals: wheat, black oat, barley, and rye. On all foods, bugs spent over 60% of their plant access time in non-probing activities. This total waveform duration was significantly longer on barley and rye compared to those on soybean and oat; wheat was intermediate. Considering only probing activities, bugs spent longer durations (ca. 2×), on soybean and oat compared to barley, rye, and wheat plants. Bugs produced significantly more pathway events on soybean and rye than on wheat and barley; with a significantly shorter duration per event on rye. The counts and durations of xylem ingestion did not differ among foods. Cell rupturing activities on seeds were longer on soybean (ca. 23%) and oat (ca. 21%), than on barley and rye (ca. 6%). The durations of ingestion events on seeds were significantly shorter on soybean (over 3×) compared to those on barley and wheat; oat and rye were intermediate. However, the ingestion duration per insect did not show significant difference among foods. Results demonstrated that D. furcatus spent more time overall in probing activities on soybean and oat; whereas, rye and barley presented the worst feeding behavior. This study provides important background information for further quantitative studies of stink bugs on different plants, such as development of resistant host plants.

Drench Application of Systemic Insecticides Disrupts Probing Behavior of Diaphorina citri (Hemiptera: Liviidae) and Inoculation of Candidatus Liberibacter asiaticus.

Candidatus Liberibacter asiaticus (CLas) is a phloem-limited bacterium that is associated with the Huanglongbing (HLB) disease of citrus and transmitted by the psyllid, Diaphorina citri. There are no curative methods to control HLB and the prevention of new infections is essential for HLB management. Therefore, the objective of our study was to determine the effects of systemic insecticides, such as the neonicotinoids imidacloprid, thiamethoxam, and a mixture of thiamethoxam and chlorantraniliprole (diamide) on the probing behavior of CLas-infected D. citri and their effect on CLas transmission. The electrical penetration graph (EPG-DC) technique was used to monitor the stylet penetration activities of CLas-infected D. citri on sweet orange [Citrus sinensis (L.) Osbeck] 'Valencia' treated with systemic insecticides. Systemic insecticides disrupted the probing behavior of CLas-infected D. citri, in a way that affected CLas transmission efficiency, particularly by negatively affecting the stylet activities related to the phloem phase. All insecticides reduced (by 57-73%) the proportion of psyllids that exhibited sustainable phloem ingestion (waveform E2 > 10 min), with significant differences observed on plants treated with thiamethoxam and thiamethoxam + chlorantraniliprole. The transmission rate of CLas with high inoculum pressure (five CLas-infected D. citri per plant and a seven-day inoculation access period) to untreated control plants was 93%. In contrast, CLas transmission was reduced to 38.8% when test plants were protected by systemic insecticides. Our results indicated that all insecticides tested presented a potential to reduce CLas inoculation by an average of 59%; therefore, these insecticides can be used to reduce the spread of HLB.

Influence of Trichomes in Strawberry Cultivars on the Feeding Behavior of Chaetosiphon fragaefolii (Cockerell) (Hemiptera: Aphididae).

Chaetosiphon fragaefolii (Cockerell) (Hemiptera: Aphididae) is the predominant aphid in strawberry (Fragaria × ananassa Duchesne) production systems in Brazil. This pest species directly damages the plants and is also responsible for spreading viruses. Further, C. fragaefolii often renders strawberry cultivation unviable, because of its high reproductive rate, as well as the large number of individuals generated through parthenogenesis. The present study aimed to (1) evaluate the feeding behavior of C. fragaefolii in four strawberry cultivars (Albion, Aromas, Camarosa, and San Andreas) and (2) identify the resistance factors associated with the number and type of trichomes in the cultivars, and also its effect on the feeding behavior of C. fragaefolii, using the electrical penetration graph (EPG) technique. The results revealed an intrinsic relationship between the number of trichomes on the cultivar and feeding behavior of C. fragaefolii. A higher number of trichomes, both tector and glandular, was observed in Albion compared to that of other cultivars, resulting in a longer no probing (Np) period per insect, and a longer Np phase. A relatively short phloem phase and ingestion time of the phloem sieve elements were also observed in Albion. These results suggest that the trichomes act as a physical barrier creating difficulties for C. fragaefolii to feed, thereby altering its feeding behavior in the four cultivars studied.

Electropenetrography (EPG): a Breakthrough Tool Unveiling Stink Bug (Pentatomidae) Feeding on Plants.

In this article, we review and discuss the potential use of EPG (electropenetrography) as a powerful tool to unveil the feeding process of phytophagous stink bugs (pentatomids). These bugs are relatively big and vigorous, which presents a problem during wiring (i.e., attachment of the gold wire on the bug's pronotum) for use in EPG. Once this challenge was overcome, using the sand paper-and-wire technique, several species have been studied using EPG, yielding waveforms that, coupled with histological studies, revealed the ingestion sites on different host plants. These sites include vascular tissues (xylem and phloem), parenchyma tissue, and seed endosperm. Stink bugs usually feed by secreting a gelling saliva to create a salivary sheath that surrounds the stylets and anchors/supports/lubricates them. However, using the cell rupture feeding strategy and the tactic of combined laceration (mechanical movements of the stylets) and maceration (action of chemical enzymes) breaks the plant cells enabling ingestion. The number of ingestion events and their duration is variable according to the feeding site. Waveforms generated have typical patterns according to the feeding site. Recent studies with several species of stink bugs have started to demonstrate the potential of EPG as a tool to unveil their feeding behavior. This may also be useful in the applied field of stink bug management, such as the development and screening of resistant genotypes and the action of chemical insecticides affecting their feeding and survivorship.

Thiamethoxam and imidacloprid drench applications on sweet orange nursery trees disrupt the feeding and settling behaviour of Diaphorina citri (Hemiptera: Liviidae).

BACKGROUND: Chemical control is the method most used for management of Diaphorina citri, the vector of the phloem-limited bacteria associated with citrus huanglongbing (HLB) disease. The objectives of this study were to determine the influence of soil-drench applications of neonicotinoids (thiamethoxam and imidacloprid) on the probing behaviour of D. citri on citrus nursery trees, using the electrical penetration graph (EPG) technique, and to measure the D. citri settling behaviour after probing on citrus nursery trees that had received these neonicotinoid treatments. RESULTS: The drench applications of neonicotinoids on citrus nursery trees disrupt D. citri probing, mainly for EPG variables related to phloem sap ingestion, with a significant reduction (≈90%) in the duration of this activity compared with untreated plants in all assessment periods (15, 35 and 90 days after application). Moreover, both insecticides have a repellent effect on D. citri, resulting in significant dispersal of psyllids from treated plants. CONCLUSIONS: This study clearly demonstrates the interference of soil-applied neonicotinoids on the feeding and settling behaviour of D. citri on citrus nursery trees, mainly during the phloem ingestion phase. These findings reinforce the recommendation of drench application of neonicotinoids before planting nursery trees as a useful strategy for HLB management. © 2015 Society of Chemical Industry.

Induced senescence promotes the feeding activities and nymph development of Myzus persicae (Hemiptera: Aphididae) on potato plants.

The effect of dark-induced senescence on Solanum tuberosum L. (Solanales: Solanaceae) plants was assessed on the feeding behavior and performance of the green peach aphid, Myzus persicae Sulzer (Hemiptera: Aphididae). Senescence was induced by covering the basal part of the plant with a black cloth for 5 d, avoiding the light passage, but keeping the apical buds uncovered. The basal part of control plants was covered with a white nonwoven cloth. The degree of senescence was determined by measuring the chlorophyll content of the covered leaves. The performance and feeding behavior of M. persicae were studied on the uncovered nonsenescent apical leaves. The aphid's performance was evaluated by measuring nymphal mortality and prereproductive time. Aphid feeding behavior was monitored by the electrical penetration graph technique. In plants with dark-induced senescence, the aphids showed a reduction in their prereproductive time. Aphids also spent more time ingesting sap from the phloem than in control plants and performed more test probes after the first sustained ingestion of phloem sap. These data suggest that M. persicae's phloem activities and nymph development benefit from the nutritional enrichment of phloem sap, derived from dark-induced senescence on potato plants. The induced senescence improved plant acceptance by M. persicae through an increase in sap ingestion that likely resulted in a reduction in developmental time.

Resistência de cultivares de batata a Myzus persicae (Sulz.)(Hemiptera: Aphididae) / Resistance of potato cultivars to Myzus persicae (Sulz.) (Hemiptera: Aphididae)

Aphids are the most important vectors of viruses infecting potato (Solanum tuberosum). We focused on the response of the aphid vector Myzus persicae (Sulzer) to five commercial potatocultivars: Ágata, Jaette Bintje, Mondial, Monalisa and Santè, by traditional antibiosis and antixenosis tests and by the EPG (Electrical Penetration Graph) technique, as a step forward to the design of effective management practices. Our aim was to identify plant factors involved in resistance of these cultivars against M. persicae, both at the surface and in deeper plant tissues. Results from the antixenosis test confirmed a strong preference of M. persicae for the Mondial cultivar. The antibiosis study indicated a lower population development of the aphid in 'Monalisa' when compared to 'Ágata' and 'Jaette Bintje'. EPG assays indicated that 'Santè' inhibited the initial feeding process of M. persicae, whereas 'Monalisa' showed a physical-type of resistance as demonstrated by a very high number of short probes. The cultivar Mondial showed average values for all EPG variables analyzed. The behavior in 'Jaette Bintje' indicated this cultivar was an ideal host for aphid feeding and reproduction. Together, the EPG data revealed the existence of pre and post-phloematics factors in the cultivars under study, which have important implications on the efficiency of transmission and spread of virus in potato by M. persicae.