Hybridisation has shaped a recent radiation of grass-feeding aphids.

BACKGROUND: Aphids are common crop pests. These insects reproduce by facultative parthenogenesis involving several rounds of clonal reproduction interspersed with an occasional sexual cycle. Furthermore, clonal aphids give birth to live young that are already pregnant. These qualities enable rapid population growth and have facilitated the colonisation of crops globally. In several cases, so-called "super clones" have come to dominate agricultural systems. However, the extent to which the sexual stage of the aphid life cycle has shaped global pest populations has remained unclear, as have the origins of successful lineages. Here, we used chromosome-scale genome assemblies to disentangle the evolution of two global pests of cereals-the English (Sitobion avenae) and Indian (Sitobion miscanthi) grain aphids. RESULTS: Genome-wide divergence between S. avenae and S. miscanthi is low. Moreover, comparison of haplotype-resolved assemblies revealed that the S. miscanthi isolate used for genome sequencing is likely a hybrid, with one of its diploid genome copies closely related to S. avenae (~ 0.5% divergence) and the other substantially more divergent (> 1%). Population genomics analyses of UK and China grain aphids showed that S. avenae and S. miscanthi are part of a cryptic species complex with many highly differentiated lineages that predate the origins of agriculture. The complex consists of hybrid lineages that display a tangled history of hybridisation and genetic introgression. CONCLUSIONS: Our analyses reveal that hybridisation has substantially contributed to grain aphid diversity, and hence, to the evolutionary potential of this important pest species. Furthermore, we propose that aphids are particularly well placed to exploit hybridisation events via the rapid propagation of live-born "frozen hybrids" via asexual reproduction, increasing the likelihood of hybrid lineage formation.

Effects of miR-306 Perturbation on Life Parameters in the English Grain Aphid, Sitobion avenae (Homoptera: Aphididae).

MicroRNAs (miRNA) play a vital role in insects' growth and development and have significant potential value in pest control. Previously, we identified miR-306 from small RNA libraries within the English grain aphid, Sitobion avenae, a devasting insect pest for wheat. miR-306 not only involves in wing morphogenesis, but also is critically important for aphid survival. Its specific impacts on the life history traits, however, remain unclear. Here, we evaluate the impact of miR-306 perturbation on S. avenae populations using a two-sex life table approach. This comprehensive analysis revealed that miR-306 perturbation significantly prolongs the developmental stages (9.64% and 8.20%) and adult longevity of S. avenae, while decreasing pre-adult survival rate (41.45% and 38.74%) and slightly reducing average fecundity (5.80% and 13.05%). Overall, miR-306 perturbation negatively affects the life table parameters of the aphid population. The population prediction models show a significant decline in the aphid population 60 days post interference, compared to the control groups (98.14% and 97.76%). Our findings highlight the detrimental effects of miR-306 perturbation on S. avenae population growth and suggest potential candidate genes for the development of RNAi-based biopesticides targeted specifically at this pest species.

Salivary Protein Cyclin-Dependent Kinase-like from Grain Aphid Sitobion avenae Suppresses Wheat Defense Response and Enhances Aphid Adaptation.

Aphids are insect pests that suck phloem sap and introduce salivary proteins into plant tissues through saliva secretion. The effector of salivary proteins plays a key role in the modulation of host plant defense responses and enhancing aphid host adaptation. Based on previous transcriptome sequencing results, a candidate effector cyclin-dependent kinase-like (CDK) was identified from the grain aphid Sitobion avenae. In this study, the function of SaCDK in wheat defense response and the adaptation of S. avenae was investigated. Our results showed that the transient overexpression of SaCDK in tobacco Nicotiana benthamiana suppressed cell death triggered by mouse pro-apoptotic protein-BAX or Phytophthora infestans PAMP-INF1. SaCDK, delivered into wheat cells through a Pseudomonas fluorescens-mediated bacterial type III secretion system, suppressed callose deposition in wheat seedlings, and the overexpression of SaCDK in wheat significantly decreased the expression levels of salicylic acid and jasmonic acid signaling pathway-related genes phenylalanine ammonia lyase (PAL), pathogenesis-related 1 protein (PR1), lipoxygenase (LOX) and Ω-3 fatty acid desaturase (FAD). In addition, aphid bioassay results showed that the survival and fecundity of S. avenae were significantly increased while feeding on the wheat plants carrying SaCDK. Taken together, our findings demonstrate that the salivary protein SaCDK is involved in inhibiting host defense response and improving its host adaptation, which lays the foundation to uncover the mechanism of the interaction of cereal aphids and host plants.

Heat-avoidance behavior associates with thermal sensitivity rather than tolerance in aphid assemblages.

How to predict animals' heat-avoidance behaviors is critical since behavior stands the first line for animals dealing with frequent heat events under ongoing climate warming. However, the discrepancy between the scarcity of research on heat-avoidance behaviors and the commonness of eco-physiological data for thermal tolerance and for thermal sensitivity such as the temperature-dependent survival time makes it difficult to link physiological thermal traits to heat-avoidance behavior. Aphids usually suck plant sap on a fixed site on the host plants at moderate temperatures, but they will leave and seek cooler feeding sites under stressful temperatures. Here we take the cereal aphid assemblages comprising different species with various development stages as a model system. We tested the hypotheses that heat tolerance (critical thermal maximum, CTmax) or heat sensitivity (temperature-dependent declining rate of survival time, similarly hereinafter) would associate with the temperature at which aphid activate heat-avoidance behavior. Specifically, we hypothesized the aphids with less heat tolerance or greater heat sensitivity would take a lower heat risk by leaving the host plant earlier. By mimicking the linear increase in ambient temperature during the daytime, we measured the CTmax and the heat-avoidance temperature (HAT, at which aphids leave the host plant to find cooler places) to understand their heat tolerance and heat-avoidance behavior. Then, we tested the survival time of aphids at different temperatures and calculated the slope of survival time declining with temperature to assess their heat sensitivity (HS). Finally, we examined the relationships between CTmax and HAT and between HS and HAT to understand if the heat-avoidance behavior associates with heat tolerance or with heat sensitivity. The results showed that HS and HAT had a strong correlation, with more heat sensitive individuals displayed lower HAT. By contrast, CTmax and HAT had a weak correlation. Our results thus provide evidence that heat sensitivity is a more reliable indicator than thermal tolerance linking with the heat-avoidance behavior in the aphid assemblages. Most existing studies use the indexes related to thermal tolerance to predict warming impacts. Our findings highlight the urgency to incorporate thermal sensitivity when predicting animal responses to climate change.

Interspecific differences in thermal tolerance landscape explain aphid community abundance under climate change.

A single critical thermal limit is often used to explain and infer the impact of climate change on geographic range and population abundance. However, it has limited application in describing the temporal dynamic and cumulative impacts of extreme temperatures. Here, we used a thermal tolerance landscape approach to address the impacts of extreme thermal events on the survival of co-existing aphid species (Metopolophium dirhodum, Sitobion avenae and Rhopalosiphum padi). Specifically, we built the thermal death time (TDT) models based on detailed survival datasets of three aphid species with three ages across a broad range of stressful high (34-40 °C) and low (-3∼-11 °C) temperatures to compare the interspecific and developmental stage variations in thermal tolerance. Using these TDT parameters, we performed a thermal risk assessment by calculating the potential daily thermal injury accumulation associated with the regional temperature variations in three wheat-growing sites along a latitude gradient. Results showed that M. dirhodum was the most vulnerable to heat but more tolerant to low temperatures than R. padi and S. avenae. R. padi survived better at high temperatures than Sitobion avenae and M. dirhodum but was sensitive to cold. R. padi was estimated to accumulate higher cold injury than the other two species during winter, while M. dirhodum accrued more heat injury during summer. The warmer site had higher risks of heat injury and the cooler site had higher risks of cold injury along a latitude gradient. These results support recent field observations that the proportion of R. padi increases with the increased frequency of heat waves. We also found that young nymphs generally had a lower thermal tolerance than old nymphs or adults. Our results provide a useful dataset and method for modelling and predicting the consequence of climate change on the population dynamics and community structure of small insects.

Wheat Oxylipins in Response to Aphids, CO2 and Nitrogen Regimes.

Wheat is critical for food security, and is challenged by biotic stresses, chiefly aphids and the viruses they transmit. The objective of this study was to determine whether aphids feeding on wheat could trigger a defensive plant reaction to oxidative stress that involved plant oxylipins. Plants were grown in chambers with a factorial combination of two nitrogen rates (100% N vs. 20% N in Hoagland solution), and two concentrations of CO2 (400 vs. 700 ppm). The seedlings were challenged with Rhopalosiphum padi or Sitobion avenae for 8 h. Wheat leaves produced phytoprostanes (PhytoPs) of the F1 series, and three types of phytofurans (PhytoFs): ent-16(RS)-13-epi-ST-Δ14-9-PhytoF, ent-16(RS)-9-epi-ST-Δ14-10-PhytoF and ent-9(RS)-12-epi-ST-Δ10-13-PhytoF. The oxylipin levels varied with aphids, but not with other experimental sources of variation. Both Rhopalosiphum padi and Sitobion avenae reduced the concentrations of ent-16(RS)-13-epi-ST-Δ14-9-PhytoF and ent-16(RS)-9-epi-ST-Δ14-10-PhytoF in relation to controls, but had little or no effect on PhytoPs. Our results are consistent with aphids affecting the levels of PUFAs (oxylipin precursors), which decreased the levels of PhytoFs in wheat leaves. Therefore, PhytoFs could be postulated as an early indicator of aphid hosting for this plant species. This is the first report on the quantification of non-enzymatic PhytoFs and PhytoPs in wheat leaves in response to aphids.

Herbivore-induced plant volatiles, not natural enemies, mediate a positive indirect interaction between insect herbivores.

Many insect herbivores engage in apparent competition whereby two species interact through shared natural enemies. Upon insect attack, plants release volatile blends that attract natural enemies, but whether these volatiles mediate apparent competition between herbivores is not yet known. We investigate the role of volatiles that are emitted by bean plants upon infestation by Acyrthosiphon pisum aphids on the population dynamics and fitness of Sitobion avenae aphids, and on wheat phloem sap metabolites. In a field experiment, the dynamics of S. avenae aphids on wheat were studied by crossing two treatments: exposure of aphid colonies to A. pisum-induced bean volatiles and exclusion of natural enemies. Glasshouse experiments and analyses of primary metabolites in wheat phloem exudates were performed to better understand the results from the field experiment. In the field, bean volatiles did not affect S. avenae dynamics or survival when aphids were exposed to natural enemies. When protected from them, however, volatiles led to larger aphid colonies. In agreement with this observation, in glasshouse experiments, aphid-induced bean volatiles increased the survival of S. avenae aphids on wheat plants, but not on an artificial diet. This suggests that volatiles may benefit S. avenae colonies via metabolic changes in wheat plants, although we did not find any effect on wheat phloem exudate composition. We report a potential case of associational susceptibility whereby plant volatiles weaken the defences of receiving plants, thus leading to increased herbivore performance.

Identification of Differentially Expressed Genes in Resistant Tetraploid Wheat (Triticum turgidum) under Sitobion avenae (F.) Infestation.

The grain aphid Sitobion avenae (Fabricius) is one of the most destructive pests of wheat (Triticum aestivum). Deployment of resistant wheat germplasm appears as an excellent solution for this problem. Elite bread wheat cultivars only have limited resistance to this pest. The present study was carried out to investigate the potential of the tetraploid wheat (Triticum turgidum) variety Lanmai, which showed high resistance to S. avenae at both seedling and adult plant stages, as a source of resistance genes. Based on apterous adult aphids' fecundity tests and choice bioassays, Lanmai has been shown to display antixenosis and antibiosis. Suppression subtractive hybridization (SSH) was employed to identify and isolate the putative candidate defense genes in Lanmai against S. avenae infestation. A total of 134 expressed sequence tags (ESTs) were identified and categorized based on their putative functions. RT-qPCR analysis of 30 selected genes confirmed their differential expression over time between the resistant wheat variety Lanmai and susceptible wheat variety Polan305 during S. avenae infestation. There were 11 genes related to the photosynthesis process, and only 3 genes showed higher expression in Lanmai than in Polan305 after S. avenae infestation. Gene expression analysis also revealed that Lanmai played a critical role in salicylic acid and jasmonic acid pathways after S. avenae infestation. This study provided further insights into the role of defense signaling networks in wheat resistance to S. avenae and indicates that the resistant tetraploid wheat variety Lanmai may provide a valuable resource for aphid tolerance improvement in wheat.

Exogenous salicylic acid improves resistance of aphid-susceptible wheat to the grain aphid, Sitobion avenae (F.) (Hemiptera: Aphididae).

Salicylic acid (SA), a phytohormone, has been considered to be a key regulator mediating plant defence against pathogens. It is still vague how SA activates plant defence against herbivores such as chewing and sucking pests. Here, we used an aphid-susceptible wheat variety to investigate Sitobion avenae response to SA-induced wheat plants, and the effects of exogenous SA on some defence enzymes and phenolics in the plant immune system. In SA-treated wheat seedlings, intrinsic rate of natural increase (rm), fecundity and apterous rate of S. avenae were 0.25, 31.4 nymphs/female and 64.4%, respectively, and significantly lower than that in the controls (P < 0.05). Moreover, the increased activities of phenylalanine-ammonia-lyase, polyphenol oxidase (PPO) and peroxidase in the SA-induced seedlings obviously depended on the sampling time, whereas activities of catalase and 4-coumarate:CoA ligase were suppressed significantly at 24, 48 and 72 h in comparison with the control. Dynamic levels of p-coumaric acid at 96 h, caffeic acid at 24 and 72 h and chlorogenic acid at 24, 48 and 96 h in wheat plants were significantly upregulated by exogenous SA application. Nevertheless, only caffeic acid content was positively correlated with PPO activity in SA-treated wheat seedlings (P = 0.031). These findings indicate that exogenous SA significantly enhanced the defence of aphid-susceptible wheat variety against aphids by regulating the plant immune system, and may prove a potential application of SA in aphid control.

Aphid fecundity and defenses in wheat exposed to a combination of heat and drought stress.

Plants are routinely subjected simultaneously to different abiotic and biotic stresses, such as heat, drought, and insect infestation. Plant-insect interactions in such complex stress situations are poorly understood. We evaluated the performance of the grain aphid (Sitobion avenae) in wheat (Triticum aestivum L.) exposed to a combination of heat and drought stresses. We also performed assays of the relative water content, nutritional quality, and responses of phytohormone signaling pathways. Lower relative water content and accumulation of soluble sugars and amino acids were observed in plants exposed to combined heat and drought stress. These conditions increased abscisic acid levels in the absence of aphids, as well as leading to higher levels of jasmonate-dependent transcripts. The grain aphid infestation further increased abscisic acid levels and the abundance of jasmonic acid- and salicylic acid-dependent defenses under the combined stress conditions. Aphids reared on plants grown under drought stress alone showed lower net reproductive rates, intrinsic rates of increase, and finite rates of increase compared with aphids reared on plants in the absence of stress. The heat-treated plants also showed a decreased aphid net reproductive rate. These findings demonstrate that exposure to a combination of stresses enhances plant defense responses against aphids as well as altering nutritional quality.

An Odorant Binding Protein (SaveOBP9) Involved in Chemoreception of the Wheat Aphid Sitobion avenae.

Odorant binding proteins play a key role in the olfactory system and are involved in the odor perception and discrimination of insects. To investigate the potential physiological functions of SaveOBP9 in Sitobion avenae, fluorescence ligand binding experiments, molecular docking, RNA interference, and behavioral tests were performed. Fluorescence binding assay results showed that SaveOBP9 had broad and high (Ki < 10 µM) binding abilities with most of the wheat volatiles, but was more obvious at pH 7.4 than pH 5.0. The binding sites of SaveOBP9 to the volatiles were predicted well by three-dimensional docking structure modeling and molecular docking. Moreover, S. avenae showed a strong behavioral response with the four compounds of wheat. The reduction in mRNA transcript levels after the RNA interference significantly reduced the expression level of SaveOBP9 and induced the non-significant response of S. avenae to the tetradecane, octanal, decanal, and hexadecane. This study provides evidence that SaveOBP9 might be involved in the chemoreception of wheat volatile organic compounds and can successfully contribute in the integrated management programs of S. avenae.

Comparative transcriptome and histological analyses of wheat in response to phytotoxic aphid Schizaphis graminum and non-phytotoxic aphid Sitobion avenae feeding.

BACKGROUND: Infestation of the phytotoxic aphid Schizaphis graminum can rapidly induce leaf chlorosis in susceptible plants, but this effect is not observed with the nonphytotoxic aphid Sitobion avenae. However, few studies have attempted to identify the different defence responses induced in wheat by S. graminum and S. avenae feeding and the mechanisms underlying the activation of chlorosis by S. graminum feeding. RESULTS: S. graminum feeding significantly reduced the chlorophyll content of wheat leaves, and these effects were not observed with S. avenae. A transcriptomic analysis showed that the expression levels of genes involved in the salicylic acid, jasmonic acid and ethylene signalling defence pathways were significantly upregulated by both S. avenae and S. graminum feeding; however, more plant defence genes were activated by S. graminum feeding than S. avenae feeding. The transcript levels of genes encoding cell wall-modifying proteins were significantly increased after S. graminum feeding, but only a few of these genes were induced by S. avenae. Furthermore, various reactive oxygen species-scavenging genes, such as 66 peroxidase (POD) and 8 ascorbate peroxidase (APx) genes, were significantly upregulated after S. graminum feeding, whereas only 15 POD and one APx genes were induced by S. avenae feeding. The activity of four antioxidant enzymes was also significantly upregulated by S. graminum feeding. Cytological examination showed that S. graminum feeding induced substantial hydrogen peroxide (H2O2) accumulation in wheat leaves. The chlorosis symptoms and the loss of chlorophyll observed in wheat leaves after S. graminum feeding were reduced and inhibited by the scavenging of H2O2 by dimethylthiourea, which indicated that H2O2 plays important role in the induction of chlorosis by S. graminum feeding. CONCLUSIONS: S. graminum and S. avenae feeding induces the JA, SA and ET signalling pathways, but S. graminum activated stronger plant defence responses than S. avenae. S. graminum feeding triggers strong ROS-scavenging activity and massive H2O2 production in wheat leaves, and the accumulation of H2O2 induced by S. graminum feeding is involved in the activation of chlorosis in wheat leaves. These results enhance our understanding of mechanisms underlying aphid-wheat interactions and provide clues for the development of aphid-resistant wheat varieties.

Defence gene expression and phloem quality contribute to mesophyll and phloem resistance to aphids in wild barley.

Aphids, including the bird cherry-oat aphid (Rhopalosiphum padi), are significant agricultural pests. The wild relative of barley, Hordeum spontaneum 5 (Hsp5), has been described to be partially resistant to R. padi, with this resistance proposed to involve higher thionin and lipoxygenase gene expression. However, the specificity of this resistance to aphids and its underlying mechanistic processes are unknown. In this study, we assessed the specificity of Hsp5 resistance to aphids and analysed differences in aphid probing and feeding behaviour on Hsp5 and a susceptible barley cultivar (Concerto). We found that partial resistance in Hsp5 to R. padi extends to two other aphid pests of grasses. Using the electrical penetration graph technique, we show that partial resistance is mediated by phloem- and mesophyll-based resistance factors that limit aphid phloem ingestion. To gain insight into plant traits responsible for partial resistance, we compared non-glandular trichome density, defence gene expression, and phloem composition of Hsp5 with those of the susceptible barley cultivar Concerto. We show that Hsp5 partial resistance involves elevated basal expression of thionin and phytohormone signalling genes, and a reduction in phloem quality. This study highlights plant traits that may contribute to broad-spectrum partial resistance to aphids in barley.

High-voltage electrostatic field-induced oxidative stress: Characterization of the physiological effects in Sitobion avenae (Hemiptera: Aphididae) across multiple generations.

In recent decades, man-made electric fields have greatly increased the intensity of electrostatic fields that are pervasively present in the environment. To better understand the physiological alterations exhibited by herbivorous insects in response to changing electric environments, we determined the activities of anti-oxidative enzymes and the metabolic rate of Sitobion avenae Fabricius (Hemiptera: Aphididae) over multiple generations in response to direct and host-seed exposure to a high-voltage electrostatic field (HVEF) of varying strength for different durations. Under controlled greenhouse conditions, 20-min direct exposure of S. avenae and wheat seeds to a 2- or 4-kV/cm HVEF resulted in significantly increased superoxide dismutase (SOD) activity in the sixth, 11th, 16th, and 21st generations relative to the control activities, whereas significantly decreased SOD activity was detected in the second generation. In addition, the activities of catalase (CAT) and peroxidase (POD) in S. avenae showed significant decreases over multiple generations. We also examined the suppressive effects of the duration of 4-kV/cm treatment on aphid physiology. The results showed that exposure to the 4-kV/cm HVEF for 20 min exerted adverse effects on CAT and POD activities and significantly decreased the metabolic rates of S. avenae, as demonstrated through evaluations of CO2 production rate, and these parameters were not significantly affected by higher HVEF durations. Overall, these findings increase our understanding of plant-pest interactions under novel HVEF environments and provide information that can improve integrated management strategies for S. avenae. Bioelectromagnetics. 40:52-61, 2019. © 2018 Wiley Periodicals, Inc.

Molecular characterization and gene silencing of Laccase 1 in the grain aphid, Sitobion avenae.

Laccase 1 (Lac1), a polyphenol oxidase, has been proposed to be involved in insect iron metabolism and immunity responses. However, little information is available on the roles of Lac 1 in insect-plant interactions. The grain aphid Sitobion avenae is one of the most destructive pests of cereal, directly drawing phloem sap and transmitting viruses. In the present study, we first cloned the open reading frame (ORF) of Lac 1 from S. avenae, and the putative protein sequence was predicted to have a carboxyl-terminal transmembrane domain. We found that SaLac1 had higher expression levels in the fourth and adult stages using reverse transcription real-time quantitative PCR (RT-qPCR). SaLac 1 was highly expressed in the salivary gland and midgut and also in wingless compared with winged morphs. After feeding on aphid-resistant wheat with a high total phenol content, the expression level of SaLac 1 increased significantly. RNA interference (RNAi) by oral feeding successfully inhibited the transcript levels of SaLac 1, and the knockdown of Lac 1 significantly decreased the survival rate of S. avenae on aphid-resistant wheat. Our study demonstrated that S. avenae Lac1 was involved in the detoxification of phenolic compounds in wheat and was essential for the aphid to adapt to resistant plants.

Searching for wheat resistance to aphids and wheat bulb fly in the historical Watkins and Gediflux wheat collections.

Insect pests can reduce wheat yield by direct feeding and transmission of plant viruses. Here we report results from laboratory and field phenotyping studies on a wide range of wheat, including landraces from the Watkins collection deriving from before the green revolution, more modern cultivars from the Gediflux collection (north-western Europe) and modern UK Elite varieties, for resistance to the bird cherry-oat aphid, Rhopalosiphum padi (Homoptera: Aphididae) and the English grain aphid, Sitobion avenae (Homoptera: Aphididae). A total of 338 lines were screened for R. padi and 340 lines for S. avenae. Field trials were also conducted on 122 Watkins lines to identify wheat bulb fly, Delia coarctata, preference on these landraces. Considerable variation was shown in insect performance among and within different wheat collections, with reduced susceptibility in a number of varieties, but phenotyping did not identify strong resistance to aphids or wheat bulb fly. Field trials showed within collection differences in aphid performance, with fewer aphids populating lines from the Watkins collection. This differs from development data in laboratory bioassays and suggests that there is a pre-alighting cue deterring aphid settlement and demonstrates differences in aphid preference and performance on older plants in the field compared with seedlings in the laboratory, highlighting the need for phenotyping for aphid resistance at different plant growth stages. No association was identified between performance of the different insect species on individual varieties, potentially suggesting different nutritional requirements or resistance mechanisms.

Deciphering the role of NADPH oxidase in complex interactions between maize (Zea mays L.) genotypes and cereal aphids.

Plant NADPH oxidases (NOXs) encompass a group of membrane-bound enzymes participating in formation of reactive oxygen species (ROS) under physiological conditions as well as in response to environmental stressors. The purpose of the survey was to unveil the role of NADPH oxidase in pro-oxidative responses of maize (Zea mays L.) seedling leaves exposed to cereal aphids' infestation. The impact of apteral females of bird cherry-oat aphid (Rhopalosiphum padi L.) and grain aphid (Sitobion avenae F.) feeding on expression levels of all four NADPH oxidase genes (rbohA, rbohB, rbohC, rbohD) and total activity of NOX enzyme in maize plants were investigated. In addition, inhibitory effect of diphenylene iodonium (DPI) pre-treatment on NOX activity and hydrogen peroxide content in aphid-stressed maize seedlings was studied. Leaf infestation biotests were accomplished on 14-day-old seedlings representing two aphid-resistant varieties (Ambrozja and Waza) and two aphid-susceptible ones (Tasty Sweet and Zlota Karlowa). Insects' attack led to profound upregulation of rbohA and rbohD genes in tested host plants, lower elevations were noted in level of rbohB mRNA, whereas abundance of rbohC transcript was not significantly altered. It was uncovered aphid-induced enhancement of NOX activity in examined plants. Higher increases in expression of all investigated rboh genes and activity of NADPH oxidase occurred in tissues of more resistant maize cultivars than in susceptible ones. Furthermore, DPI treatment resulted in strong reduction of NOX activity and H2O2 accumulation in aphid-infested Z. mays plants, thus evidencing circumstantial role of the enzyme in insect-elicited ROS generation.

SUPPRESSING A PEROXIDASE GENE REDUCES SURVIVAL IN THE WHEAT APHID Sitobion avenae.

Peroxidases (POXs) make up a large superfamily of enzymes that act in a wide range of biological mechanisms, including maintaining appropriate redox balances within cells, among other actions. In this study, we cloned a sequence that encodes a POX protein, SaPOX, from wheat aphids, Sitobion avenae. Amino acid sequence alignment showed the SaPOX sequence was conserved with POXs from other insect species. SaPOX mRNA accumulations were present in all nymphal and adult stages, at higher levels during the first and second instar, and lower during later stages in the life cycle. Ingestion of dsRNA specific to POX led to reduced SaPOX mRNA accumulation. Sitobion avenae nymphs continuously exposed to dietary dsPOX via an artificial diet led to reduced survival rate and ecdysis index. We infer that POX is important to maintain the growth and development of S. avenae.

Evidence of plastic probing behavior in a 'superclone' of the grain aphid Sitobion avenae.

Asexual reproduction is very common in invasive insect pest. In the recent years, increasing evidences have shown that some invasive asexual lineages display an outstanding capacity to predominate in space and persist on time (superclones). However, little is known about the host-use behavior of these superclones. The English grain aphid Sitobion avenae (Fabricius) (Hemiptera: Aphididae) is one of the major pests of cereals worldwide. Chilean populations of the grain aphid are characterized by a high degree of heterozygosity and low genotypic variability across regions and years, with only four predominant superclone genotypes representing nearly 90% of populations. In this study, (1) the reproductive performance and (2) the probing behavior followed a host shift of one superclone and one non-superclone of S. avenae, were compared. The host plant in the superclone did not affect the reproductive performance, while in the non-superclone was lower on highly defended wheat seedling. The experimental switching of the host plants from barley (without chemical defenses) to two wheat species with low and high levels of chemical defenses, revealed that superclone exhibited a flexible probing activities related to access of sieve elements, while the non-superclone exhibited rigid responses. These findings are consistent with the pattern of occurrence of these genotypes in the field on cereals with different plant defenses (e.g. benzoxazinoids). These responses are discussed on the view of developing new strategies for the management in invasive populations of aphid pest species.

RNA Interference of the Ecdysone Receptor Genes EcR and USP in Grain Aphid (Sitobion avenae F.) Affects Its Survival and Fecundity upon Feeding on Wheat Plants.

RNA interference (RNAi) has been widely used in functional genomics of insects and received intensive attention in the development of RNAi-based plants for insect control. Ecdysone receptor (EcR) and ultraspiracle protein (USP) play important roles in molting, metamorphosis, and reproduction of insects. EcR and USP orthologs and their function in grain aphid (Sitobion avenae F.) have not been documented yet. Here, RT-PCR, qRT-PCR, dsRNA feeding assay and aphid bioassay were employed to isolate EcR and USP orthologs in grain aphid, investigate their expression patterns, and evaluate the effect of RNAi on aphid survival and fecundity, and its persistence. The results indicated that SaEcR and SaUSP exhibited similar expression profiles at different developmental stages. Oral administration of dsRNAs of SaEcR and dsSaUSP significantly decreased the survival of aphids due to the down-regulation of these two genes, respectively. The silencing effect was persistent and transgenerational, as demonstrated by the reduced survival and fecundity due to knock-down of SaEcR and SaUSP in both the surviving aphids and their offspring, even after switching to aphid-susceptible wheat plants. Taken together, our results demonstrate that SaEcR and SaUSP are essential genes in aphid growth and development, and could be used as RNAi targets for wheat aphid control.