Search

Seemingly small ecological changes can have large, ramifying effects that defy expectations. Such are keystone effects in ecosystems. Phloem-feeding insect herbivores can act as keystone species by altering community structure and species interactions via plant-mediated or ant-mediated mechanisms. Plant responses triggered by phloem feeders can disrupt tri-trophic interactions induced by leaf-chewing herbivores, while ants that tend phloem feeders can deter or prey on other arthropods. Here, we investigate how phloem-feeding herbivores change caterpillar-parasitoid interactions on Quercus alba (white oak) trees in natural forests. We factorially manipulated the presence of phloem-feeding insects as well as ant access on Q. alba branches over multiple years and sites and measured parasitism rates of co-occurring caterpillars. While 19.3% of caterpillars were parasitized when phloem feeders were removed, the presence of phloem feeders completely suppressed parasitism of caterpillars (0%). This stark pattern was consistent across the diverse community of phloem feeders and caterpillars. Our manipulation of ant access had no effect on parasitism of caterpillars, implicating a plant-mediated mechanism. We further assessed the mechanistic hypothesis that phloem feeders suppress plant emission of caterpillar-induced volatile compounds, which could disrupt host-location behavior by parasitoids of caterpillars. Phloem feeders indeed reduced concentrations of four volatile compounds, consistent with the putative plant volatile-mediated mechanism. Given the important role of parasitoids in controlling herbivore populations, this keystone effect of phloem feeders offers novel insight into community dynamics in forests and potentially other terrestrial ecosystems., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Phloem sieve element (SE) occlusion has been hypothesized for decades to be a mechanism of resistance against phloem sap-feeding insects. Few studies have tested this hypothesis although it is likely a widespread phenomenon. This review focuses on SE occlusion by callose and P-proteins. Both are reversible, which would allow the plant to defend itself against phloem sap-feeders when SEs are penetrated and resume normal function when the insects give up and withdraw their stylets. Callose (β-1,3 glucans with some β-1,6 branches) serves many roles in plant physiology in many different tissues, each being under the control of different callose synthase genes; only callose deposited in SE sieve pores is relevant to SE occlusion. The amount of callose in sieve pores (and consequently how much it impedes sap flow) is determined by the balance in activity between callose synthase and β-1,3 glucanase. Sieve pore callose deposition has been shown to provide resistance to some phloem sap-feeders in a few studies, and in one, the difference in resistance between a susceptible and resistant rice variety was due to the ability or inability of the insect to upregulate the plants' β-1,3 glucanase that degrades the callose deposition. P-proteins occur only in dicotyledons and include a variety of proteins, not all of which are involved in SE occlusion. In some plants, P-proteins form distinct bodies in mature functional SEs. In papilionid legumes, these discrete bodies, called forisomes can expand and contract. In their expanded state, they effectively plug SEs and stop the flow of sap while in their contracted state, they provide negligible resistance to sap flow. Expansion of forisomes is triggered by an influx of Ca 2+ into the SE. Penetration of a legume (Vicia faba) SE by a generalist aphid not adapted to legumes triggers forisome expansion which occludes the SE and prevents the aphid from ingesting sap. In contrast, a legume specialist aphid, Acyrthosiphon pisum, does not trigger forisome expansion and readily ingests sap from V. faba. P-protein bodies in SEs of non-legumes do not appear to be involved in SE occlusion. In most dicotyledons, P-proteins do not form discrete bodies, but rather occur as filamentous aggregations adhering to the parietal margins of the SE and in response to damage, are released into the lumen where they are carried by the flow of sap to the downstream sieve plate where they back up and clog the sieve pores. Their effectiveness at actually stopping the flow of sap is controversial. In one study, they seemed to provide little resistance to the flow of sap while in other studies, they provided considerable resistance. In response to injury in melon, they completely stop the flow of sap, and in an aphid-resistant melon, penetration of SEs by the melon aphid, Aphis gossypii, triggers P-protein occlusion which prevents the aphids from ingesting sap. The first P-protein described, PP1, occurs only in the genus Cucurbita, and although it has been often cited to function as a SE occlusion protein, experimental evidence suggests it does not play a significant role in SE occlusion. The most common strategy for phloem sap-feeders to mitigate P-protein occlusion seems to be avoid triggering it. A widely cited in vitro study suggested that aphid saliva can reverse P-protein occlusion, but a subsequent study demonstrated that saliva was ineffective at reversing P-protein occlusion in vivo. Lastly, SE callose deposition in wheat triggered by Russian wheat aphid has been hypothesized to create an artificial sink that benefits the aphid, but additional studies are needed to test that hypothesis., (Copyright © 2021 Elsevier GmbH. All rights reserved.)

Alterations in the frequency and intensity of drought events are expected due to climate change and might have consequences for plant metabolism and the development of plant antagonists. In this study, the responses of spring wheat (Triticum aestivum) and one of its major pests, the aphid Sitobion avenae, to different drought regimes were investigated, considering different time points and plant parts. Plants were kept well-watered or subjected to either continuous or pulsed drought. Phloem exudates were collected twice from leaves and once from ears during the growth period and concentrations of amino acids, organic acids and sugars were determined. Population growth and survival of the aphid S. avenae were monitored on these plant parts. Relative concentrations of metabolites in the phloem exudates varied with the time point, the plant part as well as the irrigation regime. Pronounced increases in relative concentrations were found for proline, especially in pulsed drought-stressed plants. Moreover, relative concentrations of sucrose were lower in phloem exudates of ears than in those of leaves. The population growth and survival of aphids were decreased on plants subjected to drought and populations grew twice as large on ears compared to leaves. Our study revealed that changes in irrigation frequency and intensity modulate plant-aphid interactions. These effects may at least partly be mediated by changes in the metabolic composition of the phloem sap., Competing Interests: The authors have declared that no competing interests exist.

Woolly apple aphid (WAA), Eriosoma lanigerum (Hausmann), is an important global pest that feeds on Malus species. We studied the feeding preference of WAA on apple trees in the field for two consecutive years and in the laboratory we used electronic penetration graphs (EPG) to record the stylet penetration behavior of WAA on different parts of apple trees. We found that in the field WAA fed primarily on twigs and branches, not on leaves and fruits. Six EPG waveforms were produced during WAA probing on shoots, trunks and leaves of apple trees, including the non-penetration wave (np), the stylet pathway phase wave (C), the intracellular feeding wave (pd), the xylem feeding wave (G), waves indicative of release of saliva into the phloem (E1), and a wave indicative of ingestion from phloem (E2). In the laboratory, aphids only successfully fed on shoots, trunks and leaves, not on fruits. The EPG parameters on the phloem of shoots were significantly higher than those on trunks, indicating WAA prefer to feed on shoots. These laboratory findings explain the relative field feeding preference of WAA on different parts of apple trees, which occurs primarily on branches, barks, and young twigs in orchards, especially on young twigs., Competing Interests: The authors have declared that no competing interests exist.

To improve the proinsecticidal activity and phloem mobility of amino acid-tralopyril conjugates further, nine conjugates were designed and synthesized by introducing glutamic acid to tralopyril, and the length of the linker between glutamic acid and tralopyril ranged from 2 atoms to 10 atoms. The results of insecticidal activity against the third-instar larvae of P. xylostella showed that conjugates 42 , 43 , 44 ,and 45 (straight-chain containing 2-5 atoms) exhibited good insecticidal activity, and their LC 50 values were 0.2397 ± 0.0366, 0.4413 ± 0.0647, 0.4400 ± 0.0624, and 0.4602 ± 0.0655 mM, respectively. The concentrations of conjugates 43 - 45 were higher than that of conjugate 42 in the phloem sap at 2 h, and conjugate 43 showed the highest concentration. The introduction of glutamic acid can improve phloem mobility. The in vivo metabolism of conjugates 42 and 43 was investigated in P. xylostella , and the parent compound tralopyril was detected at concentrations of 0.5950 and 0.3172 nmol/kg, respectively. According to the above results, conjugates 42 and 43 were potential phloem mobile pro-insecticide candidates.

Background: Outbreaks of insect pests in paddy fields cause heavy losses in global rice yield annually, a threat projected to be aggravated by ongoing climate warming. Although significant progress has been made in the screening and cloning of insect resistance genes in rice germplasm and their introgression into modern cultivars, improved rice resistance is only effective against either chewing or phloem-feeding insects., Results: In this study, the results from standard and modified seedbox screening, settlement preference and honeydew excretion tests consistently showed that Qingliu, a previously known leaffolder-resistant rice variety, is also moderately resistant to brown planthopper (BPH). High-throughput RNA sequencing showed a higher number of differentially expressed genes (DEGs) at the infestation site, with 2720 DEGs in leaves vs 181 DEGs in sheaths for leaffolder herbivory and 450 DEGs in sheaths vs 212 DEGs in leaves for BPH infestation. The leaf-specific transcriptome revealed that Qingliu responds to leaffolder feeding by activating jasmonic acid biosynthesis genes and genes regulating the shikimate and phenylpropanoid pathways that are essential for the biosynthesis of salicylic acid, melatonin, flavonoids and lignin defensive compounds. The sheath-specific transcriptome revealed that Qingliu responds to BPH infestation by inducing salicylic acid-responsive genes and those controlling cellular signaling cascades. Taken together these genes could play a role in triggering defense mechanisms such as cell wall modifications and cuticular wax formation., Conclusions: This study highlighted the key defensive responses of a rarely observed rice variety Qingliu that has resistance to attacks by two different feeding guilds of herbivores. The leaffolders are leaf-feeder while the BPHs are phloem feeders, consequently Qingliu is considered to have dual resistance. Although the defense responses of Qingliu to both insect pest types appear largely dissimilar, the phenylpropanoid pathway (or more specifically phenylalanine ammonia-lyase genes) could be a convergent upstream pathway. However, this possibility requires further studies. This information is valuable for breeding programs aiming to generate broad spectrum insect resistance in rice cultivars.

On infection, plant-parasitic nematodes establish feeding sites in roots from which they take up carbohydrates among other nutrients. Knowledge on how carbohydrates are supplied to the nematodes' feeding sites is limited. Here, gene expression analyses showed that RNA levels of OsSWEET11 to OsSWEET15 were extremely low in both Meloidogyne graminicola (Mg)-caused galls and noninoculated roots. All the rice sucrose transporter genes, OsSUT1 to OsSUT5, were either down-regulated in Mg-caused galls compared with noninoculated rice roots or had very low transcript abundance. OsSUT1 was the only gene up-regulated in galls, at 14 days postinoculation (dpi), after being highly down-regulated at 3 and 7 dpi. OsSUT4 was down-regulated at 3 dpi. No noticeable OsSUTs promoter activities were detected in Mg-caused galls of pOsSUT1 to -5::GUS rice lines. Loading experiments with carboxyfluorescein diacetate (CFDA) demonstrated that symplastic connections exist between phloem and Mg-caused giant cells (GCs). According to data from OsGNS5- and OsGSL2-overexpressing rice plants that had decreased and increased callose deposition, respectively, callose negatively affected Mg parasitism and sucrose supply to Mg-caused GCs. Our results suggest that plasmodesmata-mediated sucrose transport plays a pivotal role in sucrose supply from rice root phloem to Mg-caused GCs, and OsSWEET11 to -15 and OsSUTs are not major players in it, although further functional analysis is needed for OsSUT1 and OsSUT4., (© 2021 The Authors. Molecular Plant Pathology published by British Society for Plant Pathology and John Wiley & Sons Ltd.)

Bark beetles attack their hosts at uniform intervals to avoid intraspecific competition in the phloem. Bark texture and phloem thickness also affect bark beetle attacks, and the bark characteristics are not spatially homogeneous; therefore, the distribution patterns of entry holes can demonstrate an aggregated distribution. Polygraphus proximus Blandford (Coleoptera: Scolytinae) is a non-aggressive phloephagous bark beetle that feeds on Far Eastern firs. They have caused mass mortality in Russia and Japan. However, the distribution pattern of entry holes of P. proximus and spatial relationships with bark characteristics have not been studied. Thus, we investigated the distribution pattern of entry holes of P. proximus. The distribution of entry holes was significantly uniform in most cases. As the attack density increased, an aggregated distribution pattern within a short distance (< 4.0 cm) was observed. The rough bark had a significantly higher number of entry holes than the remaining bark. The distribution pattern of entry holes demonstrated a significantly aggregated spatial association with rough bark. Finally, rough bark around knots had significantly thicker phloem than the remaining barks. These suggest that P. proximus may preferentially attack rough bark to reproduce in the thicker phloem under a rough bark surface., Competing Interests: The authors have declared that no competing interests exist.

Recent studies have reported that aphids facilitate their colonization of host plants by secreting salivary proteins into host tissues during their initial probing and feeding. Some of these salivary proteins elicit plant defenses, but the molecular and biochemical mechanisms that underlie the activation of phloem-localized resistance remain poorly understood. The aphid Myzus persicae, which is a generalized phloem-sucking pest, encompasses a number of lineages that are associated with and adapted to specific host plant species. The current study found that a cysteine protease Cathepsin B3 (CathB3), and the associated gene CathB3, was upregulated in the salivary glands and saliva of aphids from a non-tobacco-adapted (NTA) aphid lineage, when compared to those of a tobacco-adapted lineage. Furthermore, the knockdown of CathB3 improved the performance of NTA lineages on tobacco, and the propeptide domain of CathB3 was found to bind to tobacco cytoplasmic kinase ENHANCED DISEASE RESISTANCE 1-like (EDR1-like), which triggers the accumulation of reactive oxygen species in tobacco phloem, thereby suppressing both phloem feeding and colonization by NTA lineages. These findings reveal a novel function for a cathepsin-type protease in aphid saliva that elicits effective host plant defenses and warranted the theory of host specialization for generalist aphids., Competing Interests: Declaration of Interests The authors declare no competing interests., (Copyright © 2020 Elsevier Inc. All rights reserved.)

The metabolic adaptations by which phloem-feeding insects counteract plant defense compounds are poorly known. Two-component plant defenses, such as glucosinolates, consist of a glucosylated protoxin that is activated by a glycoside hydrolase upon plant damage. Phloem-feeding herbivores are not generally believed to be negatively impacted by two-component defenses due to their slender piercing-sucking mouthparts, which minimize plant damage. However, here we document that glucosinolates are indeed activated during feeding by the whitefly Bemisia tabaci. This phloem feeder was also found to detoxify the majority of the glucosinolates it ingests by the stereoselective addition of glucose moieties, which prevents hydrolytic activation of these defense compounds. Glucosylation of glucosinolates in B. tabaci was accomplished via a transglucosidation mechanism, and two glycoside hydrolase family 13 (GH13) enzymes were shown to catalyze these reactions. This detoxification reaction was also found in a range of other phloem-feeding herbivores.

The direct effects of rising global temperatures on insect herbivores could increase damage to cereal crops. However, the indirect effects of interactions between herbivores and their biotic environment at the same temperatures will potentially counter such direct effects. This study examines the potential for intraspecific competition to dampen the effects of optimal temperatures on fitness (survival × reproduction) of the brown planthopper, Nilaparvata lugens [BPH] and whitebacked planthopper, Sogatella furcifera [WBPH], two phloem-feeders that attack rice in Asia. We conducted a series of experiments with increasing densities of ovipositing females and developing nymphs on tropical and temperate rice varieties at 25, 30 and 35°C. Damage from planthoppers to the tropical variety was greater at 30°C compared to 25°C, despite faster plant growth rates at 30°C. Damage to the temperate variety from WBPH nymphs was greatest at 25°C. BPH nymphs gained greater biomass at 25°C than at 30°C despite faster development at the higher temperature (temperature-size rule); however, the effect was apparent only at high nymph densities. WBPH survival, development rates and nymph weights all declined at ≥ 30°C. At about the optimal temperature for WBPH (25°C), intraspecific crowding reduced nymph weights. Temperature has little effect on oviposition responses to density, and intraspecific competition between females only weakly counters the effects of optimal temperatures on oviposition in both BPH and WBPH. Meanwhile, the deleterious effects of nymph crowding will counter the direct effects of optimal temperatures on voltinism in BPH and on body size in both BPH and WBPH. The negative effects of crowding on BPH nymphs may be decoupled from resource use at higher temperatures., Competing Interests: The authors have declared that no competing interests exist.

Damage to grasses and cereals by phloem-feeding herbivores is manifest as nutrient and chlorophyll loss, desiccation, and a gradual decline in host vigour. Chlorophyll loss in particular leads to a succession of colour changes before eventual host death. Depending on the attacking herbivore species, colour changes can be difficult to detect with the human eye. This study used digital images to examine colour changes of rice seedlings during feeding by the brown planthopper, Nilaparvata lugens (Stål) and whitebacked planthopper, Sogatella furcifera (Horváth). Values for red (580 nm), green (540 nm) and blue (550 nm) reflectance for 39 rice varieties during seedling seed-box tests were derived from images captured with a digital camera. Red and blue reflectance gradually increased as herbivore damage progressed until final plant death. Red reflectance was greater from plants attacked by the brown planthopper than plants attacked by the whitebacked planthopper, which had proportionately more green and blue reflectance, indicating distinct impacts by the two planthoppers on their hosts. Analysis of digital images was used to discriminate variety responses to the two planthoppers. Ordination methods based on red-green-blue reflectance and vegetation indices such as the Green Leaf Index (GLI) that included blue reflectance were more successful than two-colour indices or indices based on hue, saturation and brightness in discriminating between damage responses among varieties. We make recommendations to advance seed-box screening methods for cereal resistance to phloem feeders and demonstrate how images from digital cameras can be used to improve the quality of data captured during high-throughput phenotyping., Competing Interests: The authors have declared that no competing interests exist.

The Luteoviridae are pathogenic plant viruses responsible for significant crop losses worldwide. They infect a wide range of food crops, including cereals, legumes, cucurbits, sugar beet, sugarcane, and potato and, as such, are a major threat to global food security. Viral replication is strictly limited to the plant vasculature, and this phloem limitation, coupled with the need for aphid transmission of virus particles, has made it difficult to generate virus in the quantities needed for high-resolution structural studies. Here, we exploit recent advances in heterologous expression in plants to produce sufficient quantities of virus-like particles for structural studies. We have determined their structures to high resolution by cryoelectron microscopy, providing the molecular-level insight required to rationally interrogate luteovirid capsid formation and aphid transmission, thereby providing a platform for the development of preventive agrochemicals for this important family of plant viruses., (Copyright © 2019 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Pest control in agriculture is mainly based on the application of insecticides, which may impact nontarget beneficial organisms leading to undesirable ecological effects. Neonicotinoids are among the most widely used insecticides. However, they have important negative side effects, especially for pollinators and other beneficial insects feeding on nectar. Here, we identify a more accessible exposure route: Neonicotinoids reach and kill beneficial insects that feed on the most abundant carbohydrate source for insects in agroecosystems, honeydew. Honeydew is the excretion product of phloem-feeding hemipteran insects such as aphids, mealybugs, whiteflies, and psyllids. We allowed parasitic wasps and pollinating hoverflies to feed on honeydew from hemipterans feeding on trees treated with thiamethoxam or imidacloprid, the most commonly used neonicotinoids. LC-MS/MS analyses demonstrated that both neonicotinoids were present in honeydew. Honeydew with thiamethoxam was highly toxic to both species of beneficial insects, and honeydew with imidacloprid was moderately toxic to hoverflies. Collectively, our data provide strong evidence for honeydew as a route of insecticide exposure that may cause acute or chronic deleterious effects on nontarget organisms. This route should be considered in future environmental risk assessments of neonicotinoid applications., Competing Interests: The authors declare no conflict of interest., (Copyright © 2019 the Author(s). Published by PNAS.)

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., (© The Author(s) 2019. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

The phloem is the long-distance transport system for photoassimilates within the plant. The vulnerability of the phloem tissue to blockage and damage makes it technically difficult to study, which has made it one of the least understood tissues. Transport of solution through the phloem appears to be by osmotically driven bulk flow, making an understanding of phloem hydrostatic pressure important in our comprehension of phloem flow mechanics. Here we describe a method of making in vivo direct transient measurements of phloem hydrostatic pressure using excised aphid stylets to directly access the phloem.

Background: The green peach aphid (GPA), Myzus persicae, is economically one of the most threatening pests in pepper cultivation, which not only causes direct damage but also transmits many viruses. Breeding aphid resistant pepper varieties is a promising and environmentally friendly method to control aphid populations in the field and in the greenhouse. Until now, no strong sources of resistance against the GPA have been identified. Therefore the main aims of this study were to identify pepper materials with a good level of resistance to GPA and to elucidate possible resistance mechanisms., Results: We screened 74 pepper accessions from different geographical areas for resistance to M. persicae. After four rounds of evaluation we identified one Capsicum baccatum accession (PB2013071) as highly resistant to M. persicae, while the accessions PB2013062 and PB2012022 showed intermediate resistance. The resistance of PB2013071 resulted in a severely reduced uptake of phloem compared to the susceptible accession, as determined by Electrical Penetration Graph (EPG) studies. Feeding of M. persicae induced the expression of callose synthase genes and resulted in callose deposition in the sieve elements in resistant, but not in susceptible plants., Conclusions: Three aphid resistant pepper accessions were identified, which will be important for breeding aphid resistant pepper varieties in the future. The most resistant accession PB2013071 showed phloem-based resistance against aphid infestation.

Parallel adaptations enabling the use of plant cells as the primary food source have occurred multiple times in distinct nematode clades. The hallmark of all extant obligate and facultative plant-feeding nematodes is the presence of an oral stylet, which is required for penetration of plant cell walls, delivery of pharyngeal gland secretions into host cells and selective uptake of plant assimilates. Plant parasites from different clades, and even within a single clade, display a large diversity in feeding behaviours ranging from short feeding cycles on single cells to prolonged feeding on highly sophisticated host cell complexes. Despite these differences, feeding of nematodes frequently (but certainly not always) induces common responses in host cells (e.g. endopolyploidization and cellular hypertrophy). It is thought that these host cell responses are brought about by the interplay of effectors and other biological active compounds in stylet secretions of feeding nematodes, but this has only been studied for the most advanced sedentary plant parasites. In fact, these responses are thought to be fundamental for prolonged feeding of sedentary plant parasites on host cells. However, as we discuss in this review, some of these common plant responses to independent lineages of plant parasitic nematodes might also be generic reactions to cell stress and as such their onset may not require specific inputs from plant parasitic nematodes. Sedentary plant parasitic nematodes may utilize effectors and their ability to synthesize other biologically active compounds to tailor these common responses for prolonged feeding on host cells., (© 2017 The Authors The Plant Journal © 2017 John Wiley & Sons Ltd.)

The actin cytoskeleton network has an important role in plant cell growth, division, and stress response. Actin-depolymerizing factors (ADFs) are a group of actin-binding proteins that contribute to reorganization of the actin network. Here, we show that the Arabidopsis ( Arabidopsis thaliana ) ADF3 is required in the phloem for controlling infestation by Myzus persicae Sülzer, commonly known as the green peach aphid (GPA), which is an important phloem sap-consuming pest of more than fifty plant families. In agreement with a role for the actin-depolymerizing function of ADF3 in defense against the GPA, we show that resistance in adf3 was restored by overexpression of the related ADF4 and the actin cytoskeleton destabilizers, cytochalasin D and latrunculin B. Electrical monitoring of the GPA feeding behavior indicates that the GPA stylets found sieve elements faster when feeding on the adf3 mutant compared to the wild-type plant. In addition, once they found the sieve elements, the GPA fed for a more prolonged period from sieve elements of adf3 compared to the wild-type plant. The longer feeding period correlated with an increase in fecundity and population size of the GPA and a parallel reduction in callose deposition in the adf3 mutant. The adf3 -conferred susceptibility to GPA was overcome by expression of the ADF3 coding sequence from the phloem-specific SUC2 promoter, thus confirming the importance of ADF3 function in the phloem. We further demonstrate that the ADF3 -dependent defense mechanism is linked to the transcriptional up-regulation of PHYTOALEXIN-DEFICIENT4 , which is an important regulator of defenses against the GPA., (© 2018 American Society of Plant Biologists. All Rights Reserved.)

Cucurbits have long been known to possess two types of phloem: fascicular (FP) within vascular bundles and extrafascicular phloem (EFP) surrounding vascular bundles and scattered through the cortex. Recently, their divergent composition was revealed, with FP having high sugar content consistent with conventional phloem, but EFP having much lower sugar levels and a very different proteome. However, the evolutionary advantages of possessing both FP and EFP have remained unclear. Here, we present four lines of quantitative evidence that together support the hypothesis that FP represents a typical phloem and is an attractive diet for aphids, whereas aphids avoid feeding on EFP. First, aphid stylet track endings were more abundant near the abaxial FP element of minor veins, suggesting a feeding preference for FP over EFP. Second, sugar profiles from stylet exudates were wholly consistent with FP origins, further supporting preference for FP and avoidance of EFP. Third, supplementation of EFP exudate into artificial diets confirmed an aversion to EFP in choice experiments. Finally, EFP exudate had negative effects on aphid performance. On the basis of aphids' inability to thrive on EFP, we conclude that EFP is atypical and perhaps should not be classed as a phloem system., (© 2017 John Wiley & Sons Ltd.)

The brown planthopper (BPH), Nilaparvata lugens (Stål), is a migratory and destructive sucking insect pest of rice. Silicon (Si) amendment to plants can confer enhanced resistance to herbivores and is emerging as a novel approach for pest management. In the present study, we tested the effects of Si addition at 0.16 (low) and 0.32 (high) g Si/kg soil on sucking behaviors and population growth in BPH. Si amendment increased Si content in rice stems and extended non-probing event and phloem puncture followed by sustained phloem ingestion over that in the no-Si-addition control. High Si addition rate prolonged the stylet pathway and the time needed to reach the first phloem puncture, shortened durations of phloem puncture and phloem ingestion, and decreased the proportion of individuals that produced sustained phloem ingestion. BPH female feeding on and preference for plants with the high Si addition rate were also reduced. As a result, Si application significantly decreased BPH population growth rates while increased population doubling time. These results indicate that Si amendment, especially at the high rate, confers enhanced rice plant resistance to BPH through impairment of BPH feeding. Our results highlight the potential of Si amendment as an alternative for BPH management.

Arbuscular mycorrhizal (AM) fungi are root symbionts that can increase or decrease aphid growth rates and reproduction, but the reason by which this happens is unknown. To investigate the underlying mechanisms of this interaction, we examined the effect of AM fungi on the English Grain aphid (Sitobion avenae) development, reproduction, attraction, settlement and feeding behaviour on two naturally susceptible varieties Triticum aestivum (L.) variety Solstice and T. monococcum MDR037, and two naturally resistant lines, T. monococcum MDR045 and MDR049. Mycorrhizal colonisation increased the attractiveness of T. aestivum var. Solstice to aphids, but there was no effect on aphid development on this variety. Using the Electrical Penetration Graph (EPG) technique, we found that mycorrhizal colonisation increased aphid phloem feeding on T. monococcum MDR037 and MDR045, colonisation also increased growth rate and reproductive success of S. avenae on these varieties. Mycorrhizas increased vascular bundle size, demonstrating that these fungi can influence plant anatomy. We discuss if and how this could be related to an enhanced success rate in phloem feeding in two varieties. Overall, we present and discuss how mycorrhizal fungi can affect the feeding behaviour of S. avenae in wheat, inducing susceptibility in a resistant variety.

The brown planthopper (BPH) Nilaparvata lugens is one of the most destructive insect pests on rice ( Oryza sativa ) in Asia. After landing on plants, BPH rapidly accesses plant phloem and sucks the phloem sap through unknown mechanisms. We discovered a salivary endo-β-1,4-glucanase (NlEG1) that has endoglucanase activity with a maximal activity at pH 6 at 37°C and is secreted into rice plants by BPH NlEG1 is highly expressed in the salivary glands and midgut. Silencing NlEG1 decreases the capacity of BPH to reach the phloem and reduces its food intake, mass, survival, and fecundity on rice plants. By contrast, NlEG1 silencing had only a small effect on the survival rate of BPH raised on artificial diet. Moreover, NlEG1 secreted by BPH did not elicit the production of the defense-related signal molecules salicylic acid, jasmonic acid, and jasmonoyl-isoleucine in rice, although wounding plus the application of the recombination protein NlEG1 did slightly enhance the levels of jasmonic acid and jasmonoyl-isoleucine in plants compared with the corresponding controls. These data suggest that NlEG1 enables the BPH's stylet to reach the phloem by degrading celluloses in plant cell walls, thereby functioning as an effector that overcomes the plant cell wall defense in rice., (© 2017 American Society of Plant Biologists. All Rights Reserved.)

Herbivore selection of plant hosts and plant responses to insect colonization have been subjects of intense investigations. A growing body of evidence suggests that, for successful colonization to occur, (effector/virulence) proteins in insect saliva must modulate plant defense responses to the benefit of the insect. A range of insect saliva proteins that modulate plant defense responses have been identified, but there is no direct evidence that these proteins are delivered into specific plant tissues and enter plant cells. Aphids and other sap-sucking insects of the order Hemiptera use their specialized mouthparts (stylets) to probe plant mesophyll cells until they reach the phloem cells for long-term feeding. Here, we show, by immunogold-labeling of ultrathin sections of aphid feeding sites, that an immuno-suppressive aphid effector localizes in the cytoplasm of mesophyll cells near aphid stylets but not in cells further away from aphid feeding sites. In contrast, another aphid effector protein localizes in the sheaths composed of gelling saliva that surround the aphid stylets. Thus, insects deliver effectors directly into plant tissue. Moreover, different aphid effectors locate extracellularly in the sheath saliva or are introduced into the cytoplasm of plant cells. [Formula: see text] Copyright © 2016 The Author(s). This is an open-access article distributed under the CC BY-NC-ND 4.0 International license .

The whitefly (Bemisia tabaci) causes tremendous damage to cotton production worldwide. However, very limited information is available about how plants perceive and defend themselves from this destructive pest. In this study, the transcriptomic differences between two cotton cultivars that exhibit either strong resistance (HR) or sensitivity (ZS) to whitefly were compared at different time points (0, 12, 24 and 48 h after infection) using RNA-Seq. Approximately one billion paired-end reads were obtained by Illumina sequencing technology. Gene ontology and KEGG pathway analysis indicated that the cotton transcriptional response to whitefly infestation involves genes encoding protein kinases, transcription factors, metabolite synthesis, and phytohormone signalling. Furthermore, a weighted gene co-expression network constructed from RNA-Seq datasets showed that WRKY40 and copper transport protein are hub genes that may regulate cotton defenses to whitefly infestation. Silencing GhMPK3 by virus-induced gene silencing (VIGS) resulted in suppression of the MPK-WRKY-JA and ET pathways and lead to enhanced whitefly susceptibility, suggesting that the candidate insect resistant genes identified in this RNA-Seq analysis are credible and offer significant utility. Taken together, this study provides comprehensive insights into the cotton defense system to whitefly infestation and has identified several candidate genes for control of phloem-feeding pests., (© 2016 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.)

The plant-phloem-feeding Blissus insularis possesses specialized midgut crypts, which harbor a dense population of the exocellular bacterial symbiont Burkholderia. Most individual B. insularis harbor a single Burkholderia ribotype in their midgut crypts; however, a diverse Burkholderia community exists within a host population. To understand the mechanism underlying the consistent occurrence of various Burkholderia in B. insularis and their specific association, we investigated potential gut symbiont transmission routes. PCR amplification detected a low titer of Burkholderia in adult reproductive tracts; however, fluorescence in situ hybridization assays failed to produce detectable signals in these tracts. Furthermore, no Burkholderia-specific PCR signals were detected in eggs and neonates, suggesting that it is unlikely that B. insularis prenatally transmits gut symbionts via ovarioles. In rearing experiments, most nymphs reared on St. Augustinegrass treated with cultured Burkholderia harbored the cultured Burkholderia strains. Burkholderia was detected in the untreated host grass of B. insularis, and most nymphs reared on untreated grass harbored a Burkholderia ribotype that was closely related to a plant-associated Burkholderia strain. These findings revealed that B. insularis neonates acquired Burkholderia primarily from the environment (i.e., plants and soils), even though the possibility of acquisition via egg surface cannot be excluded. In addition, our study explains how the diverse Burkholderia symbiont community in B. insularis populations can be maintained., Competing Interests: The authors have declared that no competing interests exist.

The green peach aphid, Myzus persicae Sulzer, is a notorious pest on vegetables, which often aggregates in high densities on crop leaves. In this study, we investigated whether M. persicae could suppress the resistance level of Chinese cabbage Brassica pekinensis. M. persicae performed better in terms of weight gain (~33% increase) and population growth (~110% increase) when feeding on previously infested (pre-infested) Chinese cabbage compared with those on non-infested plants. However, when given a choice, 64% of the aphids preferred to settle on non-infested leaves, while 29% of aphids chose pre-infested leaves that had a 2.9 times higher concentration of glucosinolates. Aphid feeding significantly enhanced the amino acid:sugar ratio of phloem sap and the absolute amino acid concentration in plant leaves. Aphid infestation significantly increased the expression levels of salicylic acid (SA) marker genes, while it had marginal effects on the expression of jasmonate marker genes. Exogenously applied SA or methyl jasmonate had no significant effects on M. persicae performance, although these chemicals increased glucosinolates concentration in plant leaves. M. persicae infestation increase amino acid:sugar ratio and activate plant defenses, but aphid performed better on pre-infested plants, suggesting that both nutrition and toxics should be considered in insect-plant interaction.

Increasing atmospheric carbon dioxide (CO2) impacts plant growth and metabolism. Indirectly, the performance and feeding of insects is affected by plant nutritional quality and resistance traits. Life history and feeding behaviour of Myzus persicae were studied on pepper plants under ambient (aCO2, 400 ppm) or elevated CO2 (eCO2, 650 ppm), as well as the direct impact on plant growth and leaf chemistry. Plant parameters were significantly altered by eCO2 with a negative impact on aphid's life history. Their pre-reproductive period was 11% longer and fecundity decreased by 37%. Peppers fixed significantly less nitrogen, which explains the poor aphid performance. Plants were taller and had higher biomass and canopy temperature. There was decreased aphid salivation into sieve elements, but no differences in phloem ingestion, indicating that the diminished fitness could be due to poorer tissue quality and unfavourable C:N balance, and that eCO2 was not a factor impeding feeding. Aphid ability to transmit Cucumber mosaic virus (CMV) was studied by exposing source and receptor plants to ambient (427 ppm) or elevated (612 ppm) CO2 before or after virus inoculation. A two-fold decrease on transmission was observed when receptor plants were exposed to eCO2 before aphid inoculation when compared to aCO2.

The enantiomerically enriched γ- and δ-decalactones (4a and 4b) were prepared from corresponding racemic primary-secondary 1,4- and 1,5-diols (1a and 1b), as products of enzymatic oxidation catalyzed by different alcohol dehydrogenases. The results of biotransformations indicated that the oxidation processes catalyzed by alcohol dehydrogenase (HLADH), both isolated from horse liver and recombinant in Escherichia coli, were characterized by the highest degree of conversion with moderate enantioselectivity of the reaction. Useful, environmentally friendly extraction procedure of decalactones (4a and 4b) based on hydrodistillation using a Deryng apparatus was developed. Both racemic lactones (4a and 4b), as well as their enantiomerically enriched isomers, were tested for feeding deterrent activity against Myzus persicae. The effect of these compounds on probing, feeding and settling behavior of M. persicae was studied in vivo. The deterrent activity of decalactones (4a and 4b) against aphids depended on the size of the lactone ring and the enantiomeric purity of the compounds. δ-Decalactone (4b) appeared inactive against M. persicae while γ-decalactone (4a) restrained aphid probing at ingestional phase. Only (-)-(S)-γ-decalactone (4a) had strong and durable (i.e. lasting for at least 24 hours) limiting effect, expressed at phloem level.

Signaling networks among multiple phytohormones fine-tune plant defense responses to insect herbivore attack. Previously, it was reported that the synergistic combination of ethylene (ET) and jasmonic acid (JA) was required for accumulation of the maize insect resistance1 (mir1) gene product, a cysteine (Cys) proteinase that is a key defensive protein against chewing insect pests in maize (Zea mays). However, this study suggests that mir1-mediated resistance to corn leaf aphid (CLA; Rhopalosiphum maidis), a phloem sap-sucking insect pest, is independent of JA but regulated by the ET-signaling pathway. Feeding by CLA triggers the rapid accumulation of mir1 transcripts in the resistant maize genotype, Mp708. Furthermore, Mp708 provided elevated levels of antibiosis (limits aphid population)- and antixenosis (deters aphid settling)-mediated resistance to CLA compared with B73 and Tx601 maize susceptible inbred lines. Synthetic diet aphid feeding trial bioassays with recombinant Mir1-Cys Protease demonstrates that Mir1-Cys Protease provides direct toxicity to CLA. Furthermore, foliar feeding by CLA rapidly sends defensive signal(s) to the roots that trigger belowground accumulation of the mir1, signifying a potential role of long-distance signaling in maize defense against the phloem-feeding insects. Collectively, our data indicate that ET-regulated mir1 transcript accumulation, uncoupled from JA, contributed to heightened resistance to CLA in maize. In addition, our results underscore the significance of ET acting as a central node in regulating mir1 expression to different feeding guilds of insect herbivores., (© 2015 American Society of Plant Biologists. All Rights Reserved.)

Most plant viruses that seriously damage agricultural crops are transmitted by insects. However, the mechanisms enabling virus transmission by insect vectors are poorly understood. The brown planthopper (Nilaparvata lugens) is one of the most serious rice pests, causing extensive damage to rice plants by sucking the phloem sap and transmitting viruses, including Rice ragged stunt virus (RRSV). In this study, we investigated the mechanisms of RRSV transmission from its insect vector to the rice plant in vivo using the terminal deoxynucleotidyl transferase dUTP nick-end labeling assay and RNA interference technology. RRSV induced apoptosis in the salivary gland cells of its insect vector, N. lugens. The RRSV-induced apoptosis was regulated through a caspase-dependent manner, and inhibition of the expression of N. lugens caspase-1 genes significantly interfered with virus transmission. Our findings establish a link between virus-associated apoptosis and virus transmission from the insect vector to the host plant.

Phloem tissues of pine are habitats for many thousands of organisms. Arthropods and microbes use phloem and cambium tissues to seek mates, lay eggs, rear young, feed, or hide from natural enemies or harsh environmental conditions outside of the tree. Organisms that persist within the phloem habitat are difficult to observe given their location under bark. We provide a technique to preserve intact phloem and prepare it for experimentation with invertebrates and microorganisms. The apparatus is called a 'phloem sandwich' and allows for the introduction and observation of arthropods, microbes, and other organisms. This technique has resulted in a better understanding of the feeding behaviors, life-history traits, reproduction, development, and interactions of organisms within tree phloem. The strengths of this technique include the use of inexpensive materials, variability in sandwich size, flexibility to re-open the sandwich or introduce multiple organisms through drilled holes, and the preservation and maintenance of phloem integrity. The phloem sandwich is an excellent educational tool for scientific discovery in both K-12 science courses and university research laboratories.

The Asian citrus psyllid (ACP, Diaphorina citri, Hemiptera: Liviidae), is the principal vector of the phloem-limited bacteria strongly associated with huanglongbing (HLB), the world's most serious disease of citrus. Host plant resistance may provide an environmentally safe and sustainable method of controlling ACP and/or HLB. Two xCitroncirus accessions (hybrids of Poncirus trifoliata and Citrus spp.), that are relatively resistant (UN-3881) or relatively susceptible (Troyer-1459) to ACP adults with regard to adult longevity, were compared in relation to ACP feeding behavior and some structural features of the leaf midrib. The settling (putative feeding/probing) sites of ACP adults on various parts of the leaf were not influenced primarily by plant accession. However, fewer ACP stylet sheaths were found in the midrib and fewer stylet sheath termini reached the vascular bundle (phloem and/or xylem) in UN-3881 compared to Troyer-1459 plants. Furthermore, in midribs of UN-3881 leaves the fibrous ring (sclerenchyma) around the phloem was significantly wider (thicker) compared to that in midribs of Troyer-1459 leaves. Our data indicate that feeding and/or probing by ACP adults into the vascular bundle is less frequent in the more resistant (UN-3881) than in the more susceptible (Troyer-1459) accessions. Our results also suggest that the thickness of the fibrous ring may be a barrier to stylet penetration into the vascular bundle, which is important for successful ACP feeding on the phloem and for transmitting HLB-associated bacteria. These results may help in the development of citrus plants resistant to ACP, which in turn could halt or slow the spread of the HLB-associated bacteria by this vector.

Plant viruses are known to modify the behaviour of their insect vectors, both directly and indirectly, generally adapting to each type of virus-vector relationship in a way that enhances transmission efficiency. Here, we report results of three different studies showing how a virus transmitted in a non-persistent (NP) manner (Cucumber mosaic virus; CMV, Cucumovirus) can induce changes in its host plant, cucumber (Cucumis sativus cv. Marumba) that modifies the behaviour of its aphid vector (Aphis gossypii Glover; Hemiptera: Aphididae) in a way that enhances virus transmission and spread non-viruliferous aphids changed their alighting, settling and probing behaviour activities over time when exposed to CMV-infected and mock-inoculated cucumber plants. Aphids exhibited no preference to migrate from CMV-infected to mock-inoculated plants at short time intervals (1, 10 and 30 min after release), but showed a clear shift in preference to migrate from CMV-infected to mock-inoculated plants 60 min after release. Our free-choice preference assays showed that A. gossypii alates preferred CMV-infected over mock-inoculated plants at an early stage (30 min), but this behaviour was reverted at a later stage and aphids preferred to settle and reproduce on mock-inoculated plants. The electrical penetration graph (EPG) technique revealed a sharp change in aphid probing behaviour over time when exposed to CMV-infected plants. At the beginning (first 15 min) aphid vectors dramatically increased the number of short superficial probes and intracellular punctures when exposed to CMV-infected plants. At a later stage (second hour of recording) aphids diminished their feeding on CMV-infected plants as indicated by much less time spent in phloem salivation and ingestion (E1 and E2). This particular probing behaviour including an early increase in the number of short superficial probes and intracellular punctures followed by a phloem feeding deterrence is known to enhance the transmission efficiency of viruses transmitted in a NP manner. We conclude that CMV induces specific changes in a plant host that modify the alighting, settling and probing behaviour of its main vector A. gossypii, leading to optimum transmission and spread of the virus. Our findings should be considered when modelling the spread of viruses transmitted in a NP manner., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Begomoviruses are a group of icosahedral single stranded DNA viruses exclusively transmitted by the sweet potato whitefly Bemisia tabaci in a persistent, circulative manner. In this mode of transmission, begomoviruses are acquired by their insect vector as intact virions from the plant phloem, move along the food canal, foregut and esophagus and reach the midgut where they are absorbed into the hemolymph via the filter chamber. The filter chamber is the site where most of the ingested food is filtered, and the first site where the majority of begomoviruses appear to be translocated into the hemolymph via unknown proteins or receptors. Transport from the filter chamber to the hemolymph is aided by a Heat Shock Protein 70. Virus particles not translocated across the filter chamber circulate in the midgut loop but it is not known whether absorption into the hemolymph occurs along this loop. Localization studies have confirmed that begomoviruses are not associated with the hindgut and absorption of virions in this organ is unlikely. In the hemolymph, virions have been shown to interact with a GroEL chaperone produced by the whitefly's endosymbiontic bacteria for ensuring their safe journey to the salivary glands. Virions penetrate the primary salivary glands via unknown proteins or receptors and are transported and secreted outside the whitefly to the plant with salivary secretions. Several recent studies have demonstrated the implications of insect and endosymbiont proteins such as the heat shock protein 70 and the bacterial GroEL protein, in the transmission of begomoviruses by B. tabaci. Additional studies attempting to identify other proteins that aid or interact with begomoviruses along their circulation pathway in the whitefly are reviewed in this paper., (Copyright © 2014 Elsevier B.V. All rights reserved.)

The harpin protein Hpa1 has multiple beneficial effects in plants, promoting plant growth and development, increasing crop yield, and inducing resistance to pathogens and insect pests. For these effects, the 10-40 residue fragment (Hpa1₁₀₋₄₂) isolated from the Hpa1 sequence is 1.3- to 7.5-fold more effective than the full-length protein. Here it is reported that the expression of Hpa1₁₀₋₄₂ under the direction of an insect-induced promoter induces the phloem-based defence to English grain aphid, a dominant species of wheat aphids. The expression of Hpa1₁₀₋₄₂ was found to compromise the colonization preference of aphids on the plant and further inhibit aphid reproduction in leaf colonies. In Hpa1₁₀₋₄₂-expressing wheat lines, moreover, aphid feeding from the phloem was repressed in correlation with the phloem-based defence. This defensive mechanism was shown as enhanced expression of wheat genes encoding phloem lectin proteins (PP2-A1 and PP2-A2) and β-1,3-glucan synthase-like enzymes (GSL2, GSL10, and GSL12). Both PP2-A and β-1,3-glucan formed high molecular mass polymers to block phloem sieve plate pores and therefore impede aphid feeding from the phloem. However, the phloem-based defence was impaired by treating plants with ethylene signalling inhibitors, suggesting the requirement for the ethylene signalling pathway. In addition, if Hpa1₁₀₋₄₂-expressing plants were subjected to attack by a small number of aphids, they newly acquired agriculturally beneficial characters, such as enhanced vegetative growth and increased tiller numbers and grain output values. These results suggest that the defensive and developmental roles of Hpa1₁₀₋₄₂ can be integrated into the germplasm of this agriculturally significant crop.

Sedentary endoparasitic nematodes of plants induce highly specific feeding cells in the root central cylinder. From these, the obligate parasites withdraw all required nutrients. The feeding cells were described as sink tissues in the plant's circulation system that are supplied with phloem-derived solutes such as sugars. Currently, there are several publications describing mechanisms of sugar import into the feeding cells. However, sugar processing has not been studied so far. Thus, in the present work, the roles of the sucrose-cleaving enzymes sucrose synthases (SUS) and invertases (INV) in the development of Heterodera schachtii were studied. Gene expression analyses indicate that both enzymes are regulated transcriptionally. Nematode development was enhanced on multiple INV and SUS mutants. Syncytia of these mutants were characterized by altered enzyme activity and changing sugar pool sizes. Further, the analyses revealed systemically affected sugar levels and enzyme activities in the shoots of the tested mutants, suggesting changes in the source-sink relationship. Finally, the development of the root-knot nematode Meloidogyne javanica was studied in different INV and SUS mutants and wild-type Arabidopsis plants. Similar effects on the development of both sedentary endoparasitic nematode species (root-knot and cyst nematode) were observed, suggesting a more general role of sucrose-degrading enzymes during plant-nematode interactions.

Studying plant-aphid interactions is challenging as aphid feeding is a complex process hidden in the plant tissue. Here we propose a combination of two well established methods to study nutrient acquisition by aphids focusing on the uptake of isotopically labelled nitrogen ((15)N). We combined the Electrical Penetration Graph (EPG) technique that allows detailed recording of aphid feeding behaviour and stable isotope ratio mass spectrometry (IRMS) to precisely measure the uptake of nitrogen. Bird cherry-oat aphids Rhopalosiphum padi L. (Hemiptera, Aphididae) fed for 24 h on barley plants (Hordeum vulgare L., cultivar Lina, Poaceae) that were cultivated with a (15)N enriched nutrient solution. The time aphids fed in the phloem was strongly positive correlated with their (15)N uptake. All other single behavioural phases were not correlated with (15)N enrichment in the aphids, which corroborates their classification as non-feeding EPG phases. In addition, phloem-feeding and (15)N enrichment of aphids was divided into two groups. One group spent only short time in the phloem phase and was unsuccessful in nitrogen acquisition, while the other group displayed longer phloem-feeding phases and was successful in nitrogen acquisition. This suggests that several factors such as the right feeding site, time span of feeding and individual conditions play a role for the aphids to acquire nutrients successfully. The power of this combination of methods for studying plant-aphid interactions is discussed.

There is increasing evidence that plants can react to biotic aggressions with highly specific responses. However, few studies have attempted to jointly investigate whether the induction of plant defences is specific to a targeted plant tissue, plant species, herbivore identity, and defensive trait. Here we studied those factors contributing to the specificity of induced defensive responses in two economically important pine species against two chewing insect pest herbivores. Juvenile trees of Pinus pinaster and P. radiata were exposed to herbivory by two major pest threats, the large pine weevil Hylobius abietis (a bark-feeder) and the pine processionary caterpillar Thaumetopoea pityocampa (a folivore). We quantified in two tissues (stem and needles) the constitutive (control plants) and herbivore-induced concentrations of total polyphenolics, volatile and non-volatile resin, as well as the profile of mono- and sesquiterpenes. Stem chewing by the pine weevil increased concentrations of non-volatile resin, volatile monoterpenes, and (marginally) polyphenolics in stem tissues. Weevil feeding also increased the concentration of non-volatile resin and decreased polyphenolics in the needle tissues. Folivory by the caterpillar had no major effects on needle defensive chemistry, but a strong increase in the concentration of polyphenolics in the stem. Interestingly, we found similar patterns for all these above-reported effects in both pine species. These results offer convincing evidence that induced defences are highly specific and may vary depending on the targeted plant tissue, the insect herbivore causing the damage and the considered defensive compound., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

· Small RNAs play important roles in resistance to plant viruses and the complex responses against pathogens and leaf-chewing insects. · We investigated whether small RNA pathways are involved in Arabidopsis resistance against a phloem-feeding insect, the green peach aphid (Myzus persicae). We used a 2-wk fecundity assay to assess aphid performance on Arabidopsis RNA silencing and defence pathway mutants. Quantitative real-time polymerase chain reaction was used to monitor the transcriptional activity of defence-related genes in plants of varying aphid susceptibility. High-performance liquid chromatography-mass spectrometry was employed to measure the accumulation of the antimicrobial compound camalexin. Artificial diet assays allowed the assessment of the effect of camalexin on aphid performance. · Myzus persicae produces significantly less progeny on Arabidopsis microRNA (miRNA) pathway mutants. Plants unable to process miRNAs respond to aphid infestation with increased induction of PHYTOALEXIN DEFICIENT3 (PAD3) and production of camalexin. Aphids ingest camalexin when feeding on Arabidopsis and are more successful on pad3 and cyp79b2/cyp79b3 mutants defective in camalexin production. Aphids produce less progeny on artificial diets containing camalexin. · Our data indicate that camalexin functions beyond antimicrobial defence to also include hemipteran insects. This work also highlights the extensive role of the miRNA-mediated regulation of secondary metabolic defence pathways with relevance to resistance against a hemipteran pest., (© 2013 The Authors. New Phytologist © 2013 New Phytologist Trust.)

Lens culinaris (lentil) is an important pulse crop. The yield of the crop is reduced if grown in root-knot nematode (Meloidogyne incognita) infested field. Meloidogyne incognita caused infection in primary and the secondary roots leading to the anomalies in the affected part of the root. The study revealed that the second stage juveniles (J2) of Meloidogyne incognita entered the growing roots and their branches inter and intracellularly. The immediate response was hypertrophy and hyperplasia in the root tissue near the nematode head. In response to hypertrophy some cells became very large and contained dense and granular cytoplasm. Adjacent to the giant cells, the vascular tissue was found to be disturbed. Shape, size and orientation of the vascular elements was so much altered that it had become difficult to trace the normal course of vascular strands. In various sections vascular strands were found disrupted. The vessel elements had the shapes resembling the shapes of parenchyma cells. Similarly sieve tube elements of the phloem, near the giant cells were shorter and resembled with nearby parenchyma cells. Abnormalities in xylem and phloem favored transport water, minerals and metabolites towards the giant cells. From this study, it might be inferred that alteration in the cells of galled tissue was essential for the sustenance of giant cells and for the survival of the nematode.

The soybean aphid (Aphis glycines) is a major phloem-feeding pest of soybean (Glycine max). A. glycines feeding can cause the diversion of photosynthates and transmission of plant viruses, resulting in significant yield losses. In this study, we used oligonucleotide microarrays to characterize the long-term transcriptional response to soybean aphid colonization of two related soybean cultivars, one with the Rag1 aphid-resistance gene and one aphid-susceptible cultivar (without Rag1). Transcriptome profiles were determined after 1 and 7 days of aphid infestation. Our results revealed a susceptible response involving hundreds of transcripts, whereas only one transcript changed in the resistant response to aphids. This nonexistent resistance response might be explained by the fact that many defense-related transcripts are constitutively expressed in resistant plants, whereas these same genes are activated in susceptible plants only during aphid infestation. Analysis of phytohormone-related transcripts in the susceptible response showed different hormone profiles for the two time points, and suggest that aphids are able to suppress hormone signals in susceptible plants. A significant activation of abscissic acid, normally associated with abiotic stress responses, at day 7, might be a decoy strategy implemented by the aphid to suppress effective salicylic acid- and jasmonate-related defenses.

Background: Glutathione S-transferases (GSTs) have received considerable attention in insects for their roles in insecticide resistance. Laodelphax striatellus (Fallén) is a serious rice pest. L. striatellus outbreaks occur frequently throughout eastern Asia. A key problem in controlling this pest is its rapid adaptation to numerous insecticides. In this research, nine cDNAs encoding GSTs in L. striatellus were cloned and characterised., Results: The cloned GSTs of L. striatellus belonged to six cytosolic classes and a microsomal subgroup. Exposure to sublethal concentrations of each of the six insecticides, DDT, chlorpyrifos, fipronil, imidacloprid, buprofezin and beta-cypermethrin, quickly induced (6 h) up-expression of LsGSTe1. The expression of LsGSTs2 was increased by chlorpyrifos, fipronil and beta-cypermethrin. Furthermore, exposure of L. striatellus to fipronil, imidacloprid, buprofezin and beta-cypermethrin increased the expression of the LsGSTm gene after 24 or 48 h., Conclusion: This work is the first identification of GST genes from different GST groups in Auchenorrhyncha species and their induction characteristics with insecticide types and time. The elevated expression of GST genes induced by insecticides might be related to the enhanced tolerance of this insect to insecticides and xenobiotics., (Copyright © 2012 Society of Chemical Industry.)

Background: Cowpea aphid (CPA; Aphis craccivora) is the most important insect pest of cowpea and also causes significant yield losses in other legume crops including alfalfa, beans, chickpea, lentils, lupins and peanuts. In many of these crops there is no natural genetic resistance to this sap-sucking insect or resistance genes have been overcome by newly emerged CPA biotypes., Results: In this study, we screened a subset of the Medicago truncatula core collection of the South Australian Research and Development Institute (SARDI) and identified strong resistance to CPA in a M. truncatula accession SA30199, compared to all other M. truncatula accessions tested. The biology of resistance to CPA in SA30199 plants was characterised compared to the highly susceptible accession Borung and showed that resistance occurred at the level of the phloem, required an intact plant and involved a combination of antixenosis and antibiosis. Quantitative trait loci (QTL) analysis using a F2 population (n = 150) from a cross between SA30199 and Borung revealed that resistance to CPA is controlled in part by a major quantitative trait locus (QTL) on chromosome 2, explaining 39% of the antibiosis resistance., Conclusions: The identification of strong CPA resistance in M. truncatula allows for the identification of key regulators and genes important in this model legume to give effective CPA resistance that may have relevance for other legume crops. The identified locus will also facilitate marker assisted breeding of M. truncatula for increased resistance to CPA and potentially other closely related Medicago species such as alfalfa.

Plasmodiophora brassicae (clubroot) infection leads to reprogramming of host development resulting in the formation of characteristic galls. In this work we explored the cellular events that underly gall formation in Arabidopsis thaliana with the help of molecular markers of cell division (CYCB1:GUS) and meristematic activity (ANT:GUS). Our results show that gall development involved the amplification of existing meristematic activities within the vascular cambium (VC) and phloem parenchyma (PP) cells in the region of the hypocotyl. Additionally we found that the increase in VC activity and prolonged maintenance of cambial-derived cells in a meristematic state was crucial for gall formation; disruption of the VC activity significantly decreased the gall size. Gall formation also perturbed vascular development with a significant reduction in xylem and increase in PP in infected plants. This situation was reflected in a decrease in transcripts of key factors promoting xylogenesis (VND6, VND7 and MYB46) and an increase in those promoting phloem formation and function (APL, SUC2). Finally we show, using the cell cycle inhibitor ICK1/KRP1 and a cle41 mutant with altered regulation of cambial stem cell maintenance and differentiation, that a decrease in gall formation did not prevent pathogen development. This finding demonstrates that although gall formation is a typical symptom of the disease and influences numbers of spores produced, it is not required for completion of the pathogen life cycle. Together, these results provide an insight into the relationship of the cellular events that accompany Plasmodiophora infection and their role in disease progression., (© 2012 The Authors. The Plant Journal © 2012 Blackwell Publishing Ltd.)

Sensing pH and sucrose concentration (with a preference for pH values of 7.0-7.5 and sucrose concentrations of approximately 400 mmol l(-1)) enables aphids to recognise sieve tubes inside vascular bundles. However, it is still unclear how aphids find their way to the vascular bundles. Membrane potentials in the cortex of Vicia faba stems were measured along a radial transect from the epidermis to the sieve elements and there was no gradient detected that could be used by aphids to guide their stylets to the sieve elements. Additionally, aphids did not demonstrate a preference between artificial diets with low or high levels of dissolved oxygen, making it unlikely that oxygen gradients in the cortex assist orientation towards the phloem. Tracks of salivary sheaths indicate that aphids search for vascular bundles in a radial direction (perpendicular from the stem surface to the vascular bundle) with regular side punctures in a pre-programmed fashion. Optical examination and electrical penetration graph (EPG) recordings suggest that aphids (Megoura viciae) probe the vacuolar sap of cortex cells. Acidic pH (5.0-5.5) and low sucrose concentrations in vacuoles, therefore, may provoke aphids to retract their stylets and probe the next cell until a favourable cell sap composition is encountered. The importance of sucrose as a cue was demonstrated by the experimental manipulation of Ricinus communis plants that cause them to transport hexoses instead of sucrose. Aphids (Aphis fabae) ingested less phloem sap of plants transporting hexoses compared with plants transporting the normal sucrose. The proposed rejection-acceptance behaviour provides a universal plant-directed mode of how aphids orientate their stylets towards the phloem.

Nilaparvata lugens Stål, the brown planthopper (BPH), is one of the most destructive phloem-feeding insects of rice (Oryza sativa L.) throughout Asia. Here, we show that BPH feeding increases the level of endogenous nitric oxide (NO) in the leaf and sheath tissue of both resistant and susceptible rice cultivars. However, in the roots, the NO level increased in the resistant cultivar, but decreased in the susceptible one. A burst of NO production occurred in the sheath within 1 h of infestation with BPH. The production of NO in response to BPH feeding appears to be dependent primarily on the activity of nitric oxide synthase. The application of exogenous NO reduced plant water loss by its effect on both stomatal opening and root architecture. It also stimulated the expression of certain drought stress-related genes, reduced plant height and delayed leaf senescence. Over the short term, NO supplementation reduced the seedling mortality caused by BPH feeding. This suggests that NO signaling plays a role in the rice tolerance response to BPH feeding., (Copyright © 2011 Elsevier GmbH. All rights reserved.)

Background: Pymetrozine is a valuable novel insecticide for control of sucking insects, including the brown planthopper Nilaparvata lugens (Stål), one of the most serious pests on rice. This study was conducted to elucidate the action mechanisms of pymetrozine on the feeding behaviour of the planthopper., Results: The activity test showed that pymetrozine primarily functioned as an antifeedant that caused starvation and death in N. lugens, rather than having neurotoxicity. Pymetrozine-treated insects died at a significantly slower speed than insects treated with starvation. Electrical penetration graph (EPG) data indicated that pymetrozine significantly increased the duration of non-probing periods and had a strong inhibition to phloem ingestion. The inhibition was strongly dose dependent, resulting in a complete suppression of the activity in the phloem region when the pymetrozine concentration was increased to 400 mg L(-1) . Starvation caused by inhibition of phloem ingestion might be a major toxicity mechanism of pymetrozine. EPG data also showed that pymetrozine had no significant effect on stylet movement and duration of xylem sap ingestion., Conclusion: The study revealed that pymetrozine disturbed the feeding behaviour of N. lugens mainly by increasing the non-probe period and inhibiting phloem ingestion. The inhibition resulted in a slow death similar to starvation., (Copyright © 2011 Society of Chemical Industry.)

Background: Treatment of plants with HrpNEa, a protein of harpin group produced by Gram-negative plant pathogenic bacteria, induces plant resistance to insect herbivores, including the green peach aphid Myzus persicae, a generalist phloem-feeding insect. Under attacks by phloem-feeding insects, plants defend themselves using the phloem-based defense mechanism, which is supposed to involve the phloem protein 2 (PP2), one of the most abundant proteins in the phloem sap. The purpose of this study was to obtain genetic evidence for the function of the Arabidopsis thaliana (Arabidopsis) PP2-encoding gene AtPP2-A1 in resistance to M. persicae when the plant was treated with HrpNEa and after the plant was transformed with AtPP2-A1., Results: The electrical penetration graph technique was used to visualize the phloem-feeding activities of apterous agamic M. persicae females on leaves of Arabidopsis plants treated with HrpNEa and an inactive protein control, respectively. A repression of phloem feeding was induced by HrpNEa in wild-type (WT) Arabidopsis but not in atpp2-a1/E/142, the plant mutant that had a defect in the AtPP2-A1 gene, the most HrpNEa-responsive of 30 AtPP2 genes. In WT rather than atpp2-a1/E/142, the deterrent effect of HrpNEa treatment on the phloem-feeding activity accompanied an enhancement of AtPP2-A1 expression. In PP2OETAt (AtPP2-A1-overexpression transgenic Arabidopsis thaliana) plants, abundant amounts of the AtPP2-A1 gene transcript were detected in different organs, including leaves, stems, calyces, and petals. All these organs had a deterrent effect on the phloem-feeding activity compared with the same organs of the transgenic control plant. When a large-scale aphid population was monitored for 24 hours, there was a significant decrease in the number of aphids that colonized leaves of HrpNEa-treated WT and PP2OETAt plants, respectively, compared with control plants., Conclusions: The repression in phloem-feeding activities of M. persicae as a result of AtPP2-A1 overexpression, and as a deterrent effect of HrpNEa treatment in WT Arabidopsis rather than the atpp2-a1/E/142 mutant suggest that AtPP2-A1 plays a role in plant resistance to the insect, particularly at the phloem-feeding stage. The accompanied change of aphid population in leaf colonies suggests that the function of AtPP2-A1 is related to colonization of the plant.