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Plants and insects coevolved as an evolutionarily successful and enduring association. The molecular arms race led to evolutionary novelties regarding unique mechanisms of defence and detoxification in plants and insects. While insects adopt mechanisms to conquer host defence, trees develop well-orchestrated and species-specific defence strategies against insect herbivory. However, current knowledge on the molecular underpinnings of fine-tuned tree defence responses against different herbivore insects is still restricted. In the current study, using a multi-omics approach, we unveiled the defence response of Populus tremula against aphids ( Chaitophorus populialbae ) and spongy moths ( Lymantria dispar ) herbivory. Comparative differential gene expression (DGE) analyses revealed that around 272 and 1203 transcripts were differentially regulated in P. tremula after moth and aphid herbivory compared to uninfested controls. Interestingly, 5716 transcripts were differentially regulated in P. tremula between aphids and moth infestation. Further investigation showed that defence-related stress hormones and their lipid precursors, transcription factors, and signalling molecules were over-expressed, whereas the growth-related counterparts were suppressed in P. tremula after aphid and moth herbivory. Metabolomics analysis documented that around 37% of all significantly abundant metabolites were associated with biochemical pathways related to tree growth and defence. However, the metabolic profiles of aphid and moth-fed trees were quite distinct, indicating species-specific response optimization. After identifying the suitable reference genes in P. tremula , the omics data were further validated using RT-qPCR. Nevertheless, our findings documented species-specific fine-tuning of the defence response of P. tremula, showing conservation on resource allocation for defence overgrowth under aphid and moth herbivory. Such findings can be exploited to enhance our current understanding of molecular orchestration of tree responses against herbivory and aid in developing insect pest resistance P. tremula varieties.

Insect galls are highly specialized structures arising from atypical development of plant tissue induced by insects. Galls provide the insect enhanced nutrition and protection against natural enemies and environmental stresses. Galls are essentially plant organs formed by an intimate biochemical interaction between the gall-inducing insect and its host plant. Because galls are plant organs, their development is likely to be governed by phytohormones involved in normal organogenesis. We characterized concentrations of both growth and defensive phytohormones in ungalled control leaves and galls induced by the aphid Pemphigus betae on narrowleaf cottonwood Populus angustifolia that differ genotypically in resistance to this insect. We found that susceptible trees differed from resistant trees in constitutive concentrations of both growth and defense phytohormones. Susceptible trees were characterized by significantly higher constitutive cytokinin concentrations in leaves, significantly greater ability of aphids to elicit cytokinin increases, and significantly lower constitutive defense phytohormone concentrations than observed in resistant trees. Phytohormone concentrations in both constitutive and induced responses in galled leaves exhibited high broad-sense heritability that, respectively, ranged from 0.39 to 0.93 and from 0.28 to 0.66, suggesting that selection can act upon these traits and that they might vary across the landscape. Increased cytokinin concentrations may facilitate forming strong photosynthate sinks in the galls, a requirement for galling insect success. By characterizing for the first time the changes in 15 phytohormones belonging to five different classes, this study offers a better overview of the signaling alteration occurring in galls that has likely been important for their ecology and evolution. Copyright © 2019 The Author(s). This is an open-access article distributed under the CC BY-NC-ND 4.0 International license .

Anticipated consequences of climate change in temperate regions include early spring warmup punctuated by intermittent hard freezes. Warm weather accelerates leaf flush in perennial woody species, potentially exposing vulnerable young tissues to damaging frosts. We employed a 2 × 6 randomized factorial design to examine how the interplay of vernal (springtime) freeze damage and genetic variation in a hardwood species (Populus tremuloides) influences tree growth, phytochemistry, and interactions with an insect herbivore (Chaitophorus stevensis). Acute effects of freezing included defoliation and mortality. Surviving trees exhibited reduced growth and altered biomass distribution. Reflushed leaves on these trees had lower mass per area, lower lignin concentrations, and higher nitrogen concentrations, altered chemical defence profiles, and supported faster aphid population growth. Many effects varied among plant genotypes and were related with herbivore performance. This study suggests that a single damaging vernal freeze event can alter tree-insect interactions through effects on plant growth and chemistry. Differential responses of various genotypes to freeze damage suggest that more frequent vernal freeze events could also influence natural selection, favouring trees with greater freeze hardiness, and more resistance or tolerance to herbivores following damage., (© 2017 John Wiley & Sons Ltd.)

An insect-induced gall is a highly specialized structure resulting from atypical development of plant tissue induced by a reaction to the presence and activity of an insect. The insect induces a differentiation of tissues with features and functions of an ectopic organ, providing nutrition and protection to the galling insect from natural enemies and environmental stresses. In this anatomical and cytological study, we characterized how the gall-inducing aphid Pemphigus betae reshapes the leaf morphology of the narrow-leaf cottonwood Populus angustifolia to form a leaf fold gall. Young galls displayed a bend on one side of the midvein toward the center of the leaf and back to create a fold on the abaxial side of the leaf. This fold was formed abaxially by periclinal and anticlinal divisions, effectively eliminating intercellular spaces from the spongy parenchyma. Galls at this stage exhibited both cell hypertrophy and tissue hyperplasia. Cells on the adaxial surface were more numerous and smaller than cells near the abaxial surface were, creating the large fold that surrounds the insect. Mesophyll cells exhibited some features typical of nutritive cells induced by other galling insects, including conspicuous nucleolus, reduced and fragmented vacuole, smaller and degraded chloroplasts, and dense cytoplasm compared to ungalled tissue. Even though aphids feed on the contents of phloem and do not directly consume the gall tissue, they induce changes in the plant vascular system, which lead to nutrient accumulation to support the growing aphid numbers in mature galls., Competing Interests: The authors declare that there is no conflict of interest.

The genus Populus represents one of the most economically important groups of forest trees. It is composed by approximately 30 species used for wood and non-wood products, phytoremediation and biomass. Poplar is subjected to several biological and environmental threats although, compared to annual crops, we know far less about the genetic bases of biotic stress resistance. Woolly poplar aphid (Phloeomyzus passerinii) is considered a main pest of cultivated poplars in European and American countries. In this work we present two high density linkage maps in poplar obtained by a genotyping by sequencing (GBS) approach and the identification of QTLs involved in Ph. passerinii resistance. A total of 5,667 polymorphic markers (5,606 SNPs and 61 SSRs) identified on expressed sequences have been used to genotype 131 plants of an F1 population P ×canadensis obtained by an interspecific mate between Populus deltoides (resistant to woolly poplar aphid) and Populus nigra (susceptible to woolly poplar aphid). The two linkage maps, obtained following the two-way pseudo-testcross mapping strategy, have been used to investigate the genetic bases of woolly poplar aphid resistance. One major QTL and two QTLs with minor effects (mapped on LGV, LGXVI and LG XIX) explaining the 65.8% of the genetic variance observed in the progeny in response to Ph. passerinii attack were found. The high density coverage of functional markers allowed the identification of three genes belonging to disease resistance pathway as putative candidates for P. deltoides resistance to woolly poplar aphid. This work is the first report on genetic of woolly poplar aphid genetic resistance and the resistant loci associated markers identified represent a valuable tool in resistance poplar breeding programs.

The woolly poplar aphid, Phloeomyzus passerinii (Signoret) (Hemiptera: Aphididae), is a major pest of poplar plantations in the Mediterranean basin and the Near East. Aphids colonize poplar trunks and feed upon the cortical parenchyma. Despite the economic importance of poplar, little is known about the mechanisms involved in poplar resistance to this pest. However, Populus x canadensis Moench genotypes show various levels of resistance to P. passerinii. This study has investigated the type of poplar resistance (antibiosis or antixenosis) by assessing aphid settlement, physiology (survival, development, and reproduction), and stylet penetration behavior (electrical penetration graph) on three P. x canadensis genotypes; '1214' (susceptible), 'Brenta' (resistant), and '145/51' (intermediate). Because settlement was reduced, the highly resistant genotype Brenta exhibited surface antixenosis. In addition, nymphal survival was null on Brenta, and twice less adult aphid initiated a sustained intracellular phase in the cortical parenchyma of that genotype compared with the other two genotypes. Thus, Brenta also showed parenchyma-located antixenosis coupled with antibiosis characteristic. In contrast, P. passerinii had no difficulty to initiate a sustained ingestion in the cortical parenchyma of the intermediate genotype 145/51, but decreased fecundity and lower intrinsic rate of natural increase were clear expressions of antibiosis.

We examined how the galling aphid Pemphigus batae manipulates resource translocation patterns of resistant and susceptible narrowleaf cottonwood Populus angustifolia. Using carbon-14 ((14)C)-labeling experiments in common garden trials, five patterns emerged. First, although aphid galls on resistant and susceptible genotypes did not differ in their capacity to intercept assimilates exported from the leaf they occupied, aphids sequestered 5.8-fold more assimilates from surrounding leaves on susceptible tree genotypes compared to resistant genotypes. Second, gall sinks on the same side of a shoot as a labeled leaf were 3.4-fold stronger than gall sinks on the opposite side of a shoot, which agrees with patterns of vascular connections among leaves of the same shoot (orthostichy). Third, plant genetic-based traits accounted for 26% of the variation in sink strength of gall sinks and 41% of the variation in sink strength of a plant's own bud sinks. Fourth, tree susceptibility to aphid gall formation accounted for 63% of the variation in (14)C import, suggesting strong genetic control of sink-source relationships. Fifth, competition between two galls was observed on a susceptible but not a resistant tree. On the susceptible tree distal aphids intercepted 1.5-fold more (14)C from the occupied leaf than did basal aphids, but basal aphids compensated for the presence of a distal competitor by almost doubling import to the gall from surrounding leaves. These findings and others, aimed at identifying candidate genes for resistance, argue the importance of including plant genetics in future studies of the manipulation of translocation patterns by phytophageous insects.

Genetic variation in plants is known to influence arthropod assemblages and species interactions. However, these influences may be contingent upon local environmental conditions. Here, we examine how plant genotype-based trophic interactions and patterns of natural selection change across environments. Studying the cottonwood tree, Populus angustifolia, the galling aphid, Pemphigus betae and its avian predators, we used three common gardens across an environmental gradient to examine the effects of plant genotype on gall abundance, gall size, aphid fecundity and predation rate on galls. Three patterns emerged: (i) plant genotype explained variation in gall abundance and predation, (ii) G×E explained variation in aphid fecundity, and environment explained variation in gall abundance and gall size, (iii) natural selection on gall size changed from directional to stabilizing across environments., (© 2010 The Authors. Journal of Evolutionary Biology © 2010 European Society For Evolutionary Biology.)

The aphid Chaitophorus leucomelas Koch (Hemiptera: Aphididae) is one of the most important pests of poplar (Populus spp.) plantations in Iran. In this study, development, reproduction, and life history of the aphid were assessed on 11 poplar clones; belong to three species, Populus nigra L., Populus deltoides Bartram ex Marshall, and Populus. euramericana Guinier. The experiments were carried out under controlled conditions at 24 +/- 1 degrees C, 50-60% RH, and a photoperiod of 12:12 (L:D) h. The developmental time at immature stage ranged from 10 to 12 d. Nymphs reproduced per female were ranged from 49 to 98 nymphs on Populus deltoides var. missoriensis and P. deltoides 72/51, respectively. The intrinsic rate of natural increase (r(m)) varied from 0.176 to 0.264 d(-1) among poplar clones. The r(m) values of the aphids were adversely affected in P. euramericana 242 in comparison with P. nigra 56/72 and P. nigra 63/135. In addition, the jackknife estimates of net reproductive rate (R0) indicated the presence of resistance among poplar clones. R0 ranged from 16.48 on P. nigra var. betulifoli to 47.53 on P. nigra 63/135. Mean generation times (T) was last 17.56 d on P. euramericana 242 to 14.51 d on P. deltoides 69/55. However, doubling time (DT) was 3.87 d on P. euramericana var. grandis to 2.63 d on P. nigra 63/135. The finite rate of increase (lambda) was 1.192 on resistant clone (P. euramericana 242) and 1.302 on susceptible clone (P. nigra 63/135). These results indicate that variation in life-table parameters could be an important component of variation in resistance to C. leucomelas in poplar.

We used narrowleaf cottonwood, Populus angustifolia, and the gall-forming aphid, Pemphigus betae, to determine the extent to which ontogenetic variation in resistance to herbivory is due to endogenous, stable genetic influences. In a three-year common garden trial using ramets propagated from the top, middle, and bottom of mature trees, we found that the resistance of trees to aphids was significantly higher in top vs. bottom source ramets, supporting the hypothesis of a stable, genetically programmed component to aphid resistance. The magnitude of ontogenetically based variation in resistance within an individual tree is comparable to the genetic variation in resistance among narrowleaf cottonwood genotypes or populations found in other studies. These ontogenetic-based findings have the potential to alter ecological interactions and evolutionary trajectories of plant-herbivore interactions.

Species belonging to the genus Populus (poplars) produce a series of defensive proteins in response to insect damage. Proteinase inhibitors, polyphenol oxidases, and chitinases are the most relevant and intensively studied proteins. Most of the knowledge about the relation between these proteins and herbivores has been obtained from studies with chewing insects. Nothing is known about whether phloem-feeder insects such as aphids are able to trigger a comparable response. In the current study, the expression of genes encoding a Kunitz trypsin inhibitor 3 (KTI3), a polyphenol oxidase 1 (PPO1), and a class I chitinase (CHI) was characterized in two poplar hybrids (one resistant hybrid and one susceptible hybrid, to aphids) attacked by the aphid Chaitophorus leucomelas Koch. The expression pattern was analyzed using a semiquantitative reverse transcription-polymerase chain reaction approach. The expression of KTI3 was increased by aphids only in the aphid-susceptible hybrid. Differently, PPO1 expression was increased by aphids in the aphid-resistant hybrid. The expression of CHI was down-regulated by aphids in the susceptible hybrid. This is the first study to report the differential expression of poplar defense genes in response to phloem-feeder insects such as aphids. The findings from the current study suggest that the expression levels of defensive proteins are affected by poplar genotype and by aphid infestation.

Apterous populations of Chaitophorous populicola Thomas (Homoptera: Aphididae) appear to track Eastern cottonwood (Populus deltoides Bartr.) leaf development. Few aphids occur on mature leaves. Marked individual aphids on leaves of different developmental stages were observed through a period of new leaf initiation. Nymph and adult C. populicola frequently track leaf development by moving up to younger leaves. A comparison of phloem sap constituents and leaf toughness among leaf developmental stages revealed some differences that could be used by C. populicola to determine leaf age. Phloem sap exudates, collected from P. deltoides leaves of different developmental stages, were analyzed by high-performance liquid chromatography for free amino acids and the phenolic glycoside salicin. Sucrose concentration in exudates, indicative of phloem sap exudation rate, was uniform among leaf stages. Of 20 amino acids examined, only aspartic acid and gamma-amino-n-butyric acid (GABA) concentrations differed significantly between leaf stages. Forward stepwise discriminant function analysis showed that seven of the amino acids analyzed are useful for classifying leaf maturity groupings. Aphid-infested cottonwoods had lower cystine concentrations in phloem sap than aphid-free plants. Salicin concentration was significantly higher in new leaves. Leaf toughness was assessed by lignin density and distance measurements in petiole cross-sections. Rapidly expanding leaves had significantly less lignification and new leaves had shorter distances to the vascular bundles than senescent leaves. These physiological and phytochemical differences among P. deltoides leaf developmental stages may contribute to the leaf stage selection patterns exhibited by the aphid, C. populicola.

Insects provide examples of many cunning stratagems to cope with the challenges of living in a world dominated by surface forces. Despite being the current masters of the land environment, they are at constant risk of being entrapped in liquids, which they prevent by having waxy and hairy surfaces. The problem is particularly acute in an enclosed space, such as a plant gall. Using secreted wax to efficiently parcel and transport their own excrement, aphids were able to solve this problem 200 Myr ago. Here, we report on the physical and physiological significance of this ingenious solution. The secreted powdery wax has three distinct roles: (i) it is hydrophobic, (ii) it creates a microscopically rough inner gall surface made of weakly compacted wax needles making the gall ultra-hydrophobic, and (iii) it coats the honeydew droplets converting them into liquid marbles, that can be rapidly and efficiently moved.