Jeremy Pritchard

A water-specific aquaporin involved in aphid osmoregulation.

The osmotic pressure of plant phloem sap is generally higher than that of insect body fluids. Water cycling from the distal to proximal regions of the gut is believed to contribute to the osmoregulation of aphids and other phloem-feeding insects, with the high flux of water mediated by a membrane-associated aquaporin. A putative aquaporin referred to as ApAQP1 was identified by RT-PCR of RNA isolated from the guts of pea aphids Acyrthosiphon pisum. The ApAQP1 protein has a predicted molecular mass 28.94kDa. Molecular modeling suggests that ApAQP1 has the general aquaporin topology and possesses the conserved pore properties of water-specific aquaporins. When expressed in Xenopus oocytes, ApAQP1 showed the hallmarks of aquaporin-mediated water transport, including an 18-fold increase in the osmotic water permeability of the oolemma, a reduced activation energy, and inhibition of elevated water transport activity by Hg ions. The ApAQP1 transcript was localised to the stomach and distal intestine, and RNAi-mediated knockdown of its expression resulted in elevated osmotic pressure of the haemolymph. Taken together, these data suggest that ApAQP1 contributes to the molecular basis of water cycling in the aphid gut.

Cesium toxicity in Arabidopsis

Cesium (Cs) is chemically similar to potassium (K). However, although K is an essential element, Cs is toxic to plants. Two contrasting hypotheses to explain Cs toxicity have been proposed: (1) extracellular Cs+ prevents K+ uptake and, thereby, induces K starvation; and (2) intracellular Cs+ interacts with vital K+-binding sites in proteins, either competitively or noncompetitively, impairing their activities. We tested these hypotheses with Arabidopsis (Arabidopsis thaliana). Increasing the Cs concentration in the agar ([Cs]agar) on which Arabidopsis were grown reduced shoot growth. Increasing the K concentration in the agar ([K]agar) increased the [Cs]agar at which Cs toxicity was observed. However, although increasing [Cs]agar reduced shoot K concentration ([K]shoot), the decrease in shoot growth appeared unrelated to [K]shoot per se. Furthermore, the changes in gene expression in Cs-intoxicated plants differed from those of K-starved plants, suggesting that Cs intoxication was not perceived genetically solely as K starvation. In addition to reducing [K]shoot, increasing [Cs]agar also increased shoot Cs concentration ([Cs]shoot), but shoot growth appeared unrelated to [Cs]shoot per se. The relationship between shoot growth and [Cs]shoot/[K]shoot suggested that, at a nontoxic [Cs]shoot, growth was determined by [K]shoot but that the growth of Cs-intoxicated plants was related to the [Cs]shoot/[K]shoot quotient. This is consistent with Cs intoxication resulting from competition between K+ and Cs+ for K+-binding sites on essential proteins.

A biophysical analysis of root growth under mechanical stress

The factors controlling root growth in hard soils are reviewed alongside summarised results from our recent studies. The turgor in cells in the elongation zone of roots pushes the apex forward, resisted by the external pressure of the soil and the tension in the cell walls. The external pressure of the soil consists of the pressure required to deform the soil, plus a component of frictional resistance between the root and soil. This frictional component is probably small due to the continuous sloughing of root cap cells forming a low-friction lining surrounding the root. Mechanically impeded roots are not only thicker, but are differently shaped, continuing to increase in diameter for a greater distance behind the root tip than in unimpeded roots. The osmotic potential decreases in mechanically impeded roots, possibly due to accumulation of solutes as a result of the slower root extension rate. This more negative osmotic potential is not always translated into increased turgor pressure, and the reasons for this require further investigation. The persistent effect of mechanical impedance on root growth is associated with both a stiffening of cell walls in the axial direction, and with a slowing of the rate of cell production.

High Susceptibility of Bt Maize to Aphids Enhances the Performance of Parasitoids of Lepidopteran Pests

Concerns about possible undesired environmental effects of transgenic crops have prompted numerous evaluations of such crops. So-called Bt crops receive particular attention because they carry bacteria-derived genes coding for insecticidal proteins that might negatively affect non-target arthropods. Here we show a remarkable positive effect of Bt maize on the performance of the corn leaf aphid Rhopalosiphum maidis, which in turn enhanced the performance of parasitic wasps that feed on aphid honeydew. Within five out of six pairs that were evaluated, transgenic maize lines were significantly more susceptible to aphids than their near-isogenic equivalents, with the remaining pair being equally susceptible. The aphids feed from the phloem sieve element content and analyses of this sap in selected maize lines revealed marginally, but significantly higher amino acid levels in Bt maize, which might partially explain the observed increased aphid performance. Larger colony densities of aphids on Bt plants resulted in an increased production of honeydew that can be used as food by beneficial insects. Indeed, Cotesia marginiventris, a parasitoid of lepidopteran pests, lived longer and parasitized more pest caterpillars in the presence of aphid-infested Bt maize than in the presence of aphid-infested isogenic maize. Hence, depending on aphid pest thresholds, the observed increased susceptibility of Bt maize to aphids may be either a welcome or an undesirable side effect.

Difficulties in location and acceptance of phloem sap combined with reduced concentration of phloem amino acids explain lowered performance of the aphid Rhopalosiphum padi on nitrogen deficient barley (Hordeum vulgare) seedlings.

Effects of nitrogen deficiency in hydroponically grown barley seedlings (Hordeum vulgare L.) on the development and reproduction of the aphid Rhopalosiphum padi (L.) (Hemiptera: Aphididae) were investigated.

Exploring plant responses to aphid feeding using a full Arabidopsis microarray reveals a small number of genes with significantly altered expression.

The aim of this study was to determine which Arabidopsis thaliana (L.) genes had significantly altered expression following 2-36 h of infestation by the aphid Myzus persicae (Sulzer). Six biological replicates were performed for both control and treatment at each time point, allowing rigorous statistical analysis of any changes. Only two genes showed altered expression after 2 h (one up- and one down-regulated) while two were down-regulated and twenty three were up-regulated at 36 h. The transcript annotation allowed classification of the significantly altered genes into a number of classes, including those involved in cell wall modification, carbon metabolism and signalling. Additionally, a number of genes were implicated in oxidative stress and defence against other pathogens. Five genes could not currently be assigned any function. The changes in gene expression are discussed in relation to current models of plant-insect interactions.

A mutation in amino acid permease AAP6 reduces the amino acid content of the Arabidopsis sieve elements but leaves aphid herbivores unaffected

The aim of this study was to investigate the role of the amino acid permease gene AAP6 in regulating phloem amino acid composition and then to determine the effects of this altered diet on aphid performance. A genotype of Arabidopsis thaliana (L.) was produced in which the function of the amino acid permease gene AAP6 (At5g49630) was abolished. Plants homozygous for the insertionally inactivated AAP6 gene had a significantly larger mean rosette width than the wild type and a greater number of cauline leaves. Seeds from the aap6 mutant were also significantly larger than those from the wild-type plants. Sieve element (SE) sap was collected by aphid stylectomy and the amino acids derivatized, separated, and quantified using Capillary Electrophoresis with Laser Induced Fluorescence (CE-LIF). In spite of the large variation across samples, the total amino acid concentration of SE sap of the aap6 mutant plants was significantly lower than that of the wild-type plants. The concentrations of lysine, phenylalanine, leucine, and aspartic acid were all significantly lower in concentration in the aap6 mutant plants compared with wild-type plants. This is the first direct demonstration of a physiological role for an amino acid transporter in regulating SE composition in vivo. The amino acid availability in sieve element sap is thought to be the major limiting factor for aphid growth and reproduction. Despite the changes in their diet, the aphid Myzus persicae (Sulzer) displayed only small changes in feeding behaviour on mutant plants when measured using the Electronic Penetration Graph (EPG) technique. Salivation by the aphid into the SE (E1 phase) was increased on mutant plants but there was no significant effect on other feeding EPG behaviours, or in the rate of honeydew production. Consistent with the small effect on aphid feeding behaviour, there was only a small effect of reduced sieve element amino acid concentration on aphid reproduction. The data are discussed in relation to the regulation of phloem composition and the role of phloem amino acids in regulating aphid performance.

Transpiration Induces Radial Turgor Pressure Gradients in Wheat and Maize Roots

Previous studies have shown both the presence and the absence of radial turgor and osmotic pressure gradients across the cortex of roots. In this work, gradients were sought in the roots of wheat (Triticum aestivum) and maize (Zea mays) under conditions in which transpiration flux across the root was varied This was done by altering the relative humidity above the plant, by excising the root, or by using plants in which the leaves were too young to transpire. Roots of different ages (4–65 d) were studied and radial profiles at different distances from the tip (5–30 mm) were measured. In both species, gradients of turgor and osmotic pressure (increasing inward) were found under transpiring conditions but not when transpiration was inhibited. The presence of radial turgor and osmotic pressure gradients, and the behavior of the gradient when transpiration is interrupted, indicate that active membrane transport or radial solvent drag may play an important role in the distribution of solutes across the root cortex in transpiring plants. Contrary to the conventional view, the flow of water and solutes across the symplastic pathway through the plasmodesmata cannot be inwardly directed under transpiring conditions.

Sweet problems: insect traits defining the limits to dietary sugar utilisation by the pea aphid, Acyrthosiphon pisum.

SUMMARY Plant phloem sap is an extreme diet for animals, partly because of its high and variable sugar content. The physiological and feeding traits of the pea aphid Acyrthosiphon pisum that define the upper and lower limits to the range of dietary sucrose concentrations utilised by this insect were determined principally using chemically defined diets containing 0.125–1.5 mol l–1 sucrose. On the diets with 0.125 mol l–1 and 1.5 mol l–1 sucrose, the aphids died as larvae within 8 and 14 days of birth, respectively. On the other diets, 60–96% of aphids developed to adulthood, and the 0.5 mol l–1 and 0.75 mol l–1 diets supported the highest fecundity. The diet with 0.125 mol l–1 sucrose was ingested at 36% of the rate of the 0.25 mol l–1 sucrose diet, but >90% of ingested sucrose-carbon was assimilated on both diets. This suggests that the lower limit is dictated by the aphid feeding response, specifically, a requirement for a minimal concentration of sucrose for sustained feeding. The haemolymph osmotic pressure of aphids on diets with 0.125–1.5 mol l–1 sucrose was up to 68% higher than on 0.125–1.0 mol l–1 sucrose diets, but diet consumption and sucrose-carbon assimilation was not reduced on the very high sucrose diets relative to 1.0 mol l–1 sucrose. This suggests that failure of the osmoregulatory capacity of the insects on high sucrose diets may define the upper limit to the range of dietary sucrose utilised by the aphids. The mean haemolymph osmotic pressure of aphids on plants with phloem sap containing 0.37–0.97 mol l–1 sucrose was 1.61±0.063 MPa (mean ± s.e.m.), not significantly different from that (1.47±0.059 MPa) on diets with 0.25–1.0 mol l–1 sucrose. It is concluded that the osmoregulatory response of aphids to diets and plants are comparable, and, more generally, that the feeding and osmoregulatory capabilities of the aphids are compatible with the phloem sugar levels commonly encountered by aphids feeding on plants.

A sublethal dose of thiamethoxam causes a reduction in xylem feeding by the bird cherry-oat aphid (Rhopalosiphum padi), which is associated with dehydration and reduced performance.

The active ingestion of xylem sap by aphids is hypothesised to be an important mechanism for rehydration. When starved bird cherry-oat aphids (Rhopalosiphum padi) were allowed to feed on wheat (Triticum aestivum) treated with a sublethal dose of the xylem-mobile neonicotinoid thiamethoxam, analysis of feeding behaviours using the electrical penetration graph revealed a reduction in xylem feeding that was reversed on removal of the toxin. To test the importance of xylem-feeding behaviour as a rehydration mechanism, the effects of the sublethal dose of thiamethoxam on aphid water content, honeydew excretion, growth and fecundity were investigated. Body water contents of starved R. padi feeding on wheat treated with thiamethoxam were significantly reduced compared to aphids feeding on wheat treated with distilled water (74.5+/-0.23 and 75.6+/-0.18%, respectively). In addition, the sublethal dose of thiamethoxam had detrimental effects on aphid performance. At reproductive maturity, aphids that had been born on wheat treated with thiamethoxam were significantly smaller (as measured by body plan area; 1.07+/-0.03mm(2)), lighter (0.31+/-0.04mg) and less fecund (2.85+/-0.36nymphs/day) than aphids born on wheat treated with distilled water (1.87+/-0.02mm(2), 0.72+/-0.03mg, 11.28+/-0.58nymphs/day, respectively). Regardless of whether the observed impairment of xylem feeding is due to a neurotoxic or an antifeedant effect, these results have important implications for commercial crop protection as the behaviour-modifying effects of the sublethal dose of thiamethoxam may change the efficacy of this pesticide throughout the growing season.