Immediate alteration of Macrosiphum euphorbiae host plant-selection behaviour after biotic and abiotic damage inflicted to potato plants

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1 DOI: /j x Blackwell Publishing Ltd Immediate alteration of Macrosiphum euphorbiae host plant-selection behaviour after biotic and abiotic damage inflicted to potato plants Arnaud Ameline, Aude Couty*, Sébastien Dugravot 1, Erick Campan 2, Françoise Dubois & Philippe Giordanengo Biologie des Entomophages (UPRES EA 3900), Université de Picardie Jules Verne, 33 rue St Leu, Amiens cedex, France Accepted: 14 December 2006 Key words: aphid, Homoptera, Aphididae, Solanum tuberosum, plant defence, EPG, probing behaviour, olfaction Abstract The effects of potato [Solanum tuberosum L. (Solanaceae)] plant damage on the host plant-selection behaviour of the potato aphid, Macrosiphum euphorbiae Thomas (Homoptera: Aphididae), were studied. The damage inflicted to the plant was only of short duration and observations on aphid behaviour were made immediately following plant damage. The underlying questions of the study were to know how much time it takes for plant defence mechanisms to be activated and if this activation had noticeable repercussions on aphid behaviour. We considered stresses of various natures: biotic (pre-infestation by conspecifics or by Colorado potato beetles) and abiotic (scissor cuts). Aphid responses to host plant semiochemicals were investigated using a darkened arena bioassay and the probing behaviour was assessed using the electrical penetration graph technique. Aphids were attracted to their host plant (undamaged or damaged). In a preference test (undamaged plant vs. damaged plant), plants previously infested by conspecifics were preferred to undamaged plants, but this preference was not observed for heterospecific and abiotic damage. However, aphid probing behaviour was not modified on plants previously infested by conspecifics, whereas some changes were observed subsequently to heterospecific and abiotic damages. Our data present evidence that plants can respond to biotic and abiotic stresses soon after the damage is inflicted and when the damage is of short duration. The diverse consequences of these various local plant responses on M. euphorbiae behaviour are discussed in the context of plant defence strategies against aphid colonization. Introduction Plant damage by biotic or abiotic agents may trigger induced defence mechanisms that can modify plant physiology at the physical and/or chemical level. These changes may affect behaviour and performance of phytophagous insects (Gonzales et al., 2002). The plant chemical profile can be qualitatively and/or quantitatively modified, potentially altering its quality for herbivorous insects (Karban & Baldwin, 1997; Kessler & Baldwin, *Correspondence: aude.couty@u-picardie.fr 1 Present address: Laboratoire d Ecobiologie des Insectes Parasitoïdes, UPRES EA 3193, Université Rennes I, Campus de Beaulieu, 263 Avenue du Général Leclerc, Rennes cedex, France 2 Present address: Laboratoire Dynamique de la Biodiversité (LADYBIO), UMR CNRS 5172, Université P. Sabatier Toulouse III, 118, Route de Narbonne, Toulouse cedex 04, France 2001). Moreover, in some plants, the stress-induced plant responses can be specific to the nature of the stress. Thus, chemical profiles of mechanically damaged plants often differ from those of undamaged plants and plants injured by feeding insects (Paré & Tumlinson, 1999; Walling, 2000). Injury can also induce a rapid physiological response of the plant, for instance by a sieve plate plugging mechanism, delaying phloem feeding in piercing-sucking insects (Will & van Bel, 2006). In aphids, host plant selection can be divided into several steps: host habitat location and host location are generally considered to be governed by olfactory and visual stimuli, whereas host acceptance involves olfactory, gustatory, and mechanical stimuli. Several studies have demonstrated the role of olfactory cues in host plant selection by both alate and apterous aphids (for reviews, see Pickett et al., 1992; Powell et al., 2006). Behaviour of aphids in 2007 copyright Biologie des Entomophages (UPRES EA 3900) Entomologia Experimentalis et Applicata Journal compilation 2007 The Netherlands Entomological Society 1

2 2 Ameline et al. response to odours has mainly been studied in the laboratory using olfactometers (Nottingham et al., 1991; Quiroz et al., 1997) or darkened arena bioassays (Eigenbrode et al., 2002; Jimenez-Martinez et al., 2004). Thus, attraction by host-plant odour was reported for the cotton aphid, Aphis gossypii (Pospisil, 1972), the black bean aphid, Aphis fabae (Nottingham et al., 1991), the cabbage aphid, Brevicoryne brassicae (Pettersson, 1973), and the birdcherry-oat aphid, Rhopalosiphum padi (Chamberlain et al., 2001). Modification of host plant-selection behaviour can occur following plant damage and these behavioural responses of aphids to damaged host plants can be extremely variable. Damage by pathogens such as viruses was reported to increase aphid attraction (Macias & Mink, 1969; Eigenbrode et al., 2002). Aphis fabae was attracted by the odour of its host plant but not when the leaves were mechanically damaged (Nottingham et al., 1991). Leaf damage inflicted by caterpillars repelled the lettuce aphid, Nasonovia ribis-nigri (Birkett et al., 2000), the corn leaf aphid, Rhopalosiphum maidis (Bernasconi et al., 1998), and A. fabae (Hardie et al., 1994). Attraction to a plant previously infested by conspecifics was observed for the cowpea aphid, Aphis craccivora (Pettersson et al., 1998), and the damson-hop aphid, Phorodon humuli (Campbell et al., 1993). Moreover, aphid behaviour may be affected by a density-dependent mechanism mediated by volatile compounds released at the feeding site (Pettersson et al., 1995). Aphid probing behaviour, which involves many cell punctures before reaching sieve elements, is an essential step of the host plant acceptance process (Pickett et al., 1992; Tjallingii & Hogen Esch, 1993; Prado & Tjallingii, 1999). After landing, aphids insert their stylets into epidermal cells for brief probes, using the precibarial sensilla to assess the internal plant chemistry (Harris, 1977). Therefore, plant acceptance behaviour is not directly observable as is the case for chewing insects. Electronic monitoring such as the electrical penetration graphs (EPG) technique was shown to be suitable to study probing behaviour and aphid plant relationships (McLean & Kinsey, 1967; Tjallingii, 1978, 1988). This technique, which allows analysis of all the phases belonging to the feeding behaviour of aphids, has proven to be a powerful tool for investigating the modifications of aphid feeding activities occurring on previously damaged plants. Probing behaviour was shown to be altered by a previous infestation by conspecifics (Sauge et al., 2002; Klingler et al., 2005) suggesting an induced resistance of the plant. No modification of probing behaviour was observed in R. padi after pre-infestation of Triticum aestivum by conspecifics (Prado & Tjallingii, 1997). Conversely, R. padi feeding behaviour and growth were enhanced on Sorghum halepense pre-infested by conspecifics. Pre-infestation by heterospecifics may also modify aphid feeding behaviour (Alla et al., 2001; Petersen & Sandström, 2001; Qureshi & Michaud, 2005). For example, host plant acceptance by Myzus persicae was slightly enhanced on leaves pre-infested by the heterospecific Macrosiphum euphorbiae (Thomas) (Homoptera: Aphididae) (Dugravot et al., 2007). Selection of damaged host plants can therefore be extremely variable depending on the nature of the stress affecting the host plant. However, the above-mentioned studies reported the effects of long periods of plant damage (e.g., effects of infestation initiated 4 days earlier) and long-term effects of plant damage (e.g., effects observed more than 24 h after the damage). The present experiments set out to discover if plant defence mechanisms would be triggered immediately after a short-lasting damage and if they had noticeable repercussions on aphid behaviour. Therefore, we investigated the immediate effects of biotic and abiotic damage inflicted on Solanum tuberosum L. (Solanaceae) plants, on the shortrange orientation behaviour and the feeding behaviour of apterous M. euphorbiae. The plants were damaged in various ways, and their consequences were evaluated: mechanical lesion performed with scissors (abiotic stress), feeding damage by the Colorado potato beetle, Leptinotarsa decemlineata (Say) (Coleoptera: Chrysomelidae) (heterospecific biotic stress by a chewing insect), and previous infestation by M. euphorbiae (conspecific biotic stress by a piercing/sucking insect). Materials and methods Plants and insects Solanum tuberosum var. Désirée plants were grown from tubers in 9-cm plastic pots in an environmental chamber maintained at 20 ± 1 C under a photoperiod of L16:D8. Four- to six-week-old plants were used for the experiments. Macrosiphum euphorbiae, and Colorado potato beetles (CPB) were reared on potato plants (S. tuberosum) in Plexiglas cages ( m) in environmental chambers under the same conditions used for plant growing. The M. euphorbiae colony was initiated from a single apterous parthenogenetic female from the clone Me LB05 (INRA- INSA, Villeurbanne, France). Our study focused on the apterous morph of M. euphorbiae, which accomplishes substantial interplant movements (S Dugravot, pers. obs.). The CPB colony was collected in 1999 in a field near Loos-en- Gohelle (France). Plant treatments Three types of damage were inflicted on the third leaf from the apex of the plant: (i) Abiotic damage (Ab) was performed just before the start of the bioassay

3 Immediate effects of plant damage on aphid behaviour 3 Figure 1 Drawing of the apparatus used for the darkened-arena bioassay (not to scale). (olfactometer or EPG) by cutting the leaf laterally with scissors (10 cuts, without damaging the central vein). (ii) Heterospecific damage (Hsp) was carried out using four L. decemlineata adults previously starved for 24 h. The beetles were allowed to feed for 1 h; this feeding time ensures a damaged area of 1 2 cm 2. (iii) Conspecific damage (Csp) consisted of a pre-infestation by 50 M. euphorbiae (last instar nymphs and apterous adults in random proportions) on a potato leaf inserted in an aerated plastic box ( cm). To ensure a feeding time of 1 h, in accordance with heterospecific damage, aphids were kept for 3 h with the potato leaf. Indeed, preliminary EPG experiments carried out in our experimental conditions revealed that the first phloem ingestion by M. euphorbiae on the potato plant started on average 2 h after the first contact with the plant. Insects were removed from the damaged plant just before the experiment. Undamaged potato plants (Ud) were used as control. Effect of plant damage on aphid orientation responses: set-up of dual choice behavioural experiments This bioassay was carried out in an environmentally controlled chamber at 22 ± 1 C, and 35 ± 2% r.h. in the dark to eliminate any possible visual cues. The apparatus used (Figure 1) was modified from Eigenbrode et al. (2002) and A Alvarez and WF Tjallingii (Wageningen University, unpubl.). The apparatus consisted of (i) an experimental chamber made of a Plexiglas cylinder (14 cm diameter 5 cm height) divided into two subchambers and fitted with a floor made of a double polyethylene screen (mesh size ca. 0.5 mm). Two leaflets, belonging to two distinct whole plants (or one plant leaflet and one paper leaflet), were inserted opposite each other via a 5 mm wide hole; (ii) a 9-cm Petri dish placed beneath the double screen. Fifty apterous aphids previously starved for 2 h were placed in the Petri dish at the start of the experiment. In the dark, most aphids exhibit negative geotaxis and therefore climb on the mesh (Eigenbrode et al., 2002). The double screen ensured that aphid mouthparts could not reach the leaflets. One hour after introduction, aphids positioned on either side of the mesh (corresponding to the two odour sources) were counted. Aphids located on the Petri dish were regarded as nonresponding. To avoid any possible physical bias, a rotation of 180 of the chamber was effected between replications. The chamber was cleaned with TDF4 between each replicate. Abiotically (Ab), heterospecifically (Hsp), and conspecifically (Csp) damaged potato leaves were tested vs. either an undamaged plant leaflet or a blank (paper leaflet of the same size as the potato leaflet tested). An undamaged potato leaf (Ud) was tested against a blank as a control. Seven replicates were made per treatment. For each replicate, new batches of aphids and new plants were used. Effect of plant damage on aphid probing behaviour The EPG DC system described by Tjallingii (1978, 1988) was used to investigate the probing behaviour of M. euphorbiae. In order to insert one aphid and one plant in an electrical circuit, a thin gold wire (20 µm in diameter and 2 cm long) was glued to the insect s dorsum with

4 4 Ameline et al. Table 1 Probing behaviour (mean ± SE) of Macrosiphum euphorbiae during an 8-h access to leaflets that are either Ud, undamaged (control), or Ab, abiotically (mechanically) damaged; Hsp, heterospecifically (CPB) damaged, or Csp, conspecifically (M. euphorbiae) damaged. Time and duration are expressed in minutes EPG parameters Unit Ud n = 22 Ab n = 15 Hsp n = 17 Csp n = 17 General probing behaviour 1 Number of probes # 14.8 ± ± ± ± Total duration of probing min ± ± ± ± Time from start of recording to first probe min 17.0 ± ± ± ± 5.3 Pathway phase 4 Number of pathway phases # 20.5 ± ± ± ± Total duration of pathway phases min ± ± ± ± 16.3 Salivation phase 6 Number of single salivation phases # 2.7 ± ± ± 0.3* 2.9 ± Total duration of single salivation phases min 5.9 ± ± ± 1.3* 5.9 ± Time from start to first salivation min 84.5 ± ± ± 20.9* 93.3 ± 22.8 Phloem ingestion 9 Total time of phloem ingestion min ± ± ± ± Total time of sustained phloem ingestion min ± ± ± ± Time from start to first phloem ingestion min ± ± ± 27.6* ± Time from start to first sustained phloem ingestion min ± ± 37.0* ± 29.3* ± Number of probes before the first phloem ingestion # 7.5 ± ± 2.7* 11.4 ± 1.2* 11.9 ± Number of probes before the first sustained phloem ingestion # 10.4 ± ± 2.8* 14.4 ± 1.5* 16.1 ± 2.6 Other parameters 15 Total time of stylet derailment min 33.0 ± ± ± ± Total time of xylem ingestion min 2.6 ± ± ± ± 7.5 Means followed by * indicate a significant difference with control plants according to Mann Whitney U-test at P<0.05. conductive silver paint and the other electrode was inserted into the soil of the potted plant. All plants and insects were held inside a Faraday cage during recording at an ambient temperature of 20 ± 1 C. The recordings were performed with one aphid per plant continuously for 8 h during the day. Two- to three-day-old adult apterous females were used. The acquisition and analysis of the EPG waveforms were done with PROBE 3.0 software (WF Tjallingii, Wageningen University, The Netherlands). Macrosiphum euphorbiae probing behaviour was monitored on the plant leaflets damaged as described above (Ab, Hsp, or Csp) and compared with undamaged control plants (Ud). For each treatment, between 16 and 22 replicates were done. Sixteen parameters based on the five different EPG waveforms (C, E1, E2, F, and G) described by Tjallingii (1978, 1988) were calculated using the EPG Calc visual basic macro (P Giordanengo, unpubl.): C waveform corresponding to stylet pathways in all tissues except phloem and xylem, E1 to salivation in phloem elements, E2 to passive phloem sap ingestion, G to active xylem sap ingestion, and F to stylet penetration difficulties (termed stylet derailment ). The 16 EPG parameters were assigned to five categories (Table 1): (i) general probing behaviour class: this included the mean number of probes by the aphids, the total duration of probing, and the time spent by aphids before initiation of the first probe; (ii) pathway phases that corresponded to searching activities performed by the aphids before reaching phloem elements; this class included the number of pathway phases and their total duration; (iii) salivation phases, with single salivation occurring when a salivation period into phloem vessels was not followed by sap ingestion, and the time from start of recording to first salivation period corresponded to the time spent by aphids before reaching the phloem elements; (iv) phloem ingestion phase (distinction was made between a phloem ingestion period shorter than10 min and a sustained phloem ingestion period longer than 10 min, considered as a real ingestion phase); (v) other EPG parameters were attributed to xylem ingestion and to stylet derailment. Statistical analyses Statistical analyses were performed using STATISTICA 5.5 software (StatSoft, Tulsa, OK, USA). A Kruskall Wallis

5 Immediate effects of plant damage on aphid behaviour 5 Figure 2 Distribution of responding apterous Macrosiphum euphorbiae in the darkened bioassay arena. The bars represent the percentage of responding aphids that made a particular choice; the total number (n) is given on the left side of each experimental situation. Ab, abiotically (mechanically) damaged potato leaflet; Hsp, heterospecifically (CPB) damaged potato leaflet; and Csp, conspecifically (M. euphorbiae) damaged potato leaflet were tested vs. either an Ud (undamaged potato leaflet) or a blank. As a control, an undamaged potato leaf (Ud) vs. a blank was tested. Asterisks indicate a significant difference (P<0.05) in the distribution of aphids between the two sides of the darkened arena. n.s., not significant, at α = ANOVA was performed to test the effect of treatment (Ab, Hsp, Csp, and Ud) on the number of non-responding insects in the darkened arena bioassay. Results are presented as percentages. For each treatment in the dual choice experiments, the distribution of the responding aphids on either side of the mesh was analysed using a Wilcoxon test for paired samples. As data were not normally distributed, the 16 EPG parameters (cf. Table 1) of the damaged plants (Ab, Hsp, and Csp) were compared pairwise with control plants (Ud) by non-parametric Mann Whitney U-test at P = Results Aphid orientation response to damaged potato plants The mean percentage of non-responding insects ranged from 21 to 57 and was not significantly different between the experimental situations (Kruskall Wallis: H = 10.25, P = 0.17); therefore, we only considered the responding aphids for further analysis. Results are expressed in percentages in Figure 2. When tested against a blank, M. euphorbiae was significantly attracted by the potato leaflet whether undamaged (Ud: 69%, Wilcoxon: Z = 2.197, P = 0.028) or damaged (Ab: 73%, Wilcoxon: Z = 2.201, P = 0.028; Hsp: 68%, Wilcoxon, Z = and P = 0.018; Csp: 69%, Wilcoxon: Z = 2.197, P = 0.028). Aphids were significantly more attracted by an undamaged plant than by a scissordamaged plant (Ud: 64% vs. Ab: 36%, Wilcoxon: Z = 2.028, P = 0.042) while no preference was expressed between an undamaged plant (Ud) and a CPB (Hsp)-damaged plant (Wilcoxon: Z = 0.33, P = 0.73). In contrast, plants conspecifically damaged were significantly preferred to undamaged ones (Csp: 61% vs. Ud: 39%, Wilcoxon: Z = 2.366, P = 0.018). Aphid feeding behaviour on damaged potato plants Results are presented in Table 1. Parameters related to phloem ingestion were significantly modified when aphids fed on an Ab-damaged plant. The time spent by aphids before performing the first sustained phloem ingestion period and the number of probes preceding first phloem ingestion, sustained or not, were increased (Mann Whitney U-tests for parameters 12 14: U = 120.5, P = 0.027; U = 115, P = 0.005; and U = 97, P = 0.019, respectively). When aphid probing behaviour was monitored on a leaflet of a Hsp-damaged plant, the same alterations as reported for the Ab treatment were observed on parameters related to phloem ingestion (Mann Whitney U-tests for parameters 12 14: U = 119.5, P = 0.020; U = 121, P = 0.022; U = 134.5, P = 0.009, respectively). Moreover, the first phloem ingestion was also delayed (Mann Whitney U-tests for parameter 11: U = 103 and P = 0.008). On these Hspdamaged plants, all the EPG parameters related to the salivation phase were also modified: the number of single salivations and their total duration were significantly reduced and the first salivation was delayed (Mann Whitney U-tests for parameters 6 and 7: U = 114.5, P = and U = 144, P = 0.040, respectively). Parameters in relation to general probing behaviour and pathway phases, and also the total time of stylet derailment and xylem ingestion, were not modified whatever the nature of the damage (Ab, Csp, or Hsp) inflicted to the plant. When the feeding behaviour of M. euphorbiae was monitored on a Csp leaflet, none of the EPG parameter values were different from those of aphids feeding on undamaged plants.

6 6 Ameline et al. Discussion The role of olfaction in aphid host-plant location has been well documented (for a review, see Powell et al., 2006), but most studies have focused on the colonizing morphs (i.e., the winged morphs) showing their attraction towards their host plant. We report here that apterous M. euphorbiae are attracted by potato plant volatiles. Such attraction of apterous aphids towards their host plant has been previously described (Nottingham et al., 1991; Bernasconi et al., 1998; Alla et al., 2003). The novelty of this work, however, lies in the demonstration of an immediate alteration of aphid host plant selection behaviour following damage inflicted to a plant over a short period of time (1 h). Although previous studies had focused on olfaction (Quiroz et al., 1997; Bernasconi et al., 1998) and feeding behaviour (Prado & Tjallingii, 1997; Sauge et al., 2002), they were conducted after a substantially longer period of plant damage to evaluate long-term effects on aphids. Our study shows that various sorts of stress, including abiotic, heterospecific, or conspecific, can immediately affect not only aphid orientation behaviour but also its feeding behaviour. Effect of heterospecific and abiotic damage on aphid behaviour Both CPB- and scissor-damaged potato leaves triggered aphid orientation when offered in a dual choice test against a blank. This is apparently not in accordance with the work of Nottingham et al. (1991) who showed that A. fabae was attracted by undamaged Vicia faba plants but not by a mechanically damaged plant. In addition, this same aphid species was repelled by a host plant chewed by a caterpillar (Hardie et al., 1994). Such a repellency of a host plant induced by a chewing insect has been reported for several aphid species (Bernasconi et al., 1998; Paré & Tumlinson, 1999). It is interesting to note that repellency or nonattractiveness of mechanically damaged plants was shown by these authors after a long period (at least 24 h after the damage) whereas in our study, aphid behaviour was observed 1 h after the damage. It is therefore possible that potentially repulsive semiochemicals had not yet been induced in our experimental conditions. Although abiotically and heterospecifically damaged plants were attractive when tested against a blank, they were not preferred to an undamaged plant in a dual-choice situation. Indeed, aphids preferred an undamaged potato plant to an abiotically scissor-damaged one and they expressed no preference between a CPB-damaged plant and an undamaged potato plant. Kruzmane et al. (2002) showed that CPB regurgitant on mechanically wounded potato led to specific induction of plant defence signalling cascades. Thus, the observed divergence in aphid preferences could be due to differences between the chemical profiles of CPB-damaged and scissor-damaged plants. Variation related to the nature of the injury inflicted to the plant was also observed on the feeding behaviour of M. euphorbiae. When monitored on an abiotically damaged plant, time to first sustained phloem ingestion period was delayed, suggesting either a chemically or physically induced plant defence. It can be hypothesized that this delay was linked to a lower phloem appetency due to modification of the chemical composition of the phloem sap or to a lower phloem accessibility due to physical changes in the phloem tubes (Will & van Bel, 2006). Concerning the phloem ingestion parameters, the same alterations as previously described were observed when aphids fed on CPB-damaged leaves. In addition, both first brief phloem ingestion (parameter 11) and first salivation (parameter 8) were significantly delayed. This supports the hypothesis of an alteration of either plant physical or chemical properties. Conversely, the reduced number and duration of single salivation phases are usually associated with an aphid feeding improvement (Prado & Tjallingii, 1997; Sauge et al., 2002). This apparent contradiction leads us to suppose that, whereas phloem vessels are less accessible, phloem sap is easier to ingest from a damaged plant than from an undamaged one. The occurrence of a putative elicitor in CPB saliva (Kruzmane et al., 2002) could affect aphid salivation (frequency and duration). Effect of conspecific damage on aphid behaviour Macrosiphum euphorbiae oriented preferentially towards a leaflet previously infested by conspecifics rather than towards an intact plant. Previous studies have shown that the presence of an aphid colony on a plant can cause arrestment or aggregation of conspecifics of A. fabae (Kay, 1976) and A. craccivora (Pettersson et al., 1998). In our experiments, although aphids were removed from the plant just before the test, we could not rule out that traces of some aphid-emitted compounds persisted on the leaflet surface. Another hypothesis would be the release of plant semiochemicals specifically induced by aphids (Chamberlain et al., 2001). Such aphid-induced plant responses have also been reported to affect aphid feeding behaviour. For instance, Sauge et al. (2002) showed a modification of M. persicae probing behaviour on plants previously infested by conspecifics. This alteration of behaviour depended on the susceptibility of the peach cultivar used: a pre-infestation on a resistant cultivar induced a higher plant resistance whereas a pre-infestation on a susceptible cultivar induced facilitation. A facilitation of feeding behaviour after conspecific pre-infestation was also reported for A. fabae on its Vicia faba host plant but not for R. padi on its Triticum

7 Immediate effects of plant damage on aphid behaviour 7 aestivum host plant (Prado & Tjallingii, 1997). This shows the diversity of plant/aphid interactions, which could be linked to the complexity of the variation in gene expression induced by aphid infestation in various plants (Voelckel et al., 2004; Zhu-Salzman et al., 2004). In our case, M. euphorbiae probing behaviour was not modified on a potato plant previously infested by conspecifics. This lack of feeding behaviour modification could be linked to the short duration of the pre-infestation period: 1 h vs. the 2 5 days pre-infestation in the other studies (Prado & Tjallingii, 1997; Sauge et al., 2002). Ecological consequences Our data show that induced plant responses, leading either to beneficial or detrimental effects on aphid colonization processes, can occur very soon after plant damage and that aphids are capable of detecting this immediate local plant response. In the context of our study (immediate effect of damage of short duration), abiotic mechanical injury on potato foliage reduced plant attractiveness towards aphids whereas a CPB injury had no effect. More durable mechanical damage, whether biotic or abiotic, has been shown to induce significant emission of terpenes by plants several hours after damage. Such terpenes include β-farnesene, which is repellent towards aphids but also attracts aphid natural enemies (Pickett et al., 1992). On the contrary, short-duration damage by conspecifics were responsible for aphid short-range attraction. More intense and more prolonged aphid damage could affect not only plant phytochemistry and aphid feeding behaviour (Gianoli & Niemeyer, 1997) but also aphid fitness (Wool & Hales, 1996; Qureshi & Michaud, 2005). Indeed, changes in plant phytochemistry subsequent to the injection of aphid saliva in the phloem elements (Prado & Tjallingii, 1994) generally occur in favour of the aphid s own nutritional benefit (Petersen & Sandström, 2001). In addition, Tosh et al. (2002) showed that larviposition is only initiated after phloem acceptance and ingestion. These data suggest that potato pre-infestation by aphids could lead to subsequent facilitation of colonization by conspecifics. It is also worthwhile to note that in terms of plant phytochemical responses, herbivore feeding is not equivalent to mechanical wounding. Indeed, different plant defence-signalling pathways are activated depending on the nature of the injury, whether mechanical or through herbivore feeding. In addition, among herbivorous insects, distinction has to be made between piercing/sucking insects such as aphids, that induce pathways similar to those triggered by pathogens (salicylic acid-dependent and jasmonic acid/ethylene-dependent pathways) and chewing insects that induce wound-signalling pathways (mainly jasmonic acid-dependent pathway) (Walling, 2000). Our work focused on immediate effects of various kinds of plant damage on aphid behaviour. We also only considered the local plant response as we only tested the leaflet that was damaged. Further studies are needed to investigate whether similar effects can be observed in the longer term and also if the plant response is spread to the whole plant by systemic signalling. Such information would be of considerable interest for the general understanding of the temporal dynamics of aphid plant interactions in a complex agro-ecosystem where plants can be challenged, often simultaneously, by a number of herbivores. Acknowledgements The work was supported by the Ministère Français de la Recherche, the Conseil Régional de Picardie, and the Comité Nord Plants de Pommes de Terre. We thank Y. Rahbé (INRA, Villeurbanne, France) for providing the M. euphorbiae clone and the Comité Nord Plants de Pommes de Terre for providing the potato tubers. We also wish to thank Andrew Roots for his critical reading of the manuscript. References Alla S, Moreau JP & Frérot B (2001) Effects on the aphid Rhopalosiphum padi on the leafhopper Psamotettix alienus under laboratory conditions. Entomologia Experimentalis et Applicata 98: Alla S, Cherqui A, Kaiser L, Azzouz H, Sangwan-Norreel BS & Giordanengo P (2003) Effects of potato plants expressing the nptii-gus fusion marker genes on reproduction, longevity, and host-finding of the peach potato aphid, Myzus persicae. Entomologia Experimentalis et Applicata 106: Bernasconi M, Turlings TCJ, Ambrosetti L, Bassetti P & Dorn S (1998) Herbivore-induced emissions of maize volatiles repel the corn leaf aphid, shape Rhopalosiphum maidis. Entomologia Experimentalis et Applicata 87: Birkett MA, Campbell CAM, Chamberlain K, Guerrieri E, Hick AJ et al. (2000) New roles for cis-jasmone as an insect semiochemical and in plant defense. Proceedings of the National Academy of Sciences, USA 97: Campbell CAM, Pettersson J, Pickett JA, Wadhams L & Woodcock C (1993) Spring migration of damson-hop aphid, Phorodon humuli (Homoptera, Aphididae), on summer host plant-derived semiochemicals released on feeding. Journal of Chemical Ecology 19: Chamberlain K, Guerrieri E, Pennacchio F, Pettersson J, Pickett JA et al. (2001) Can aphid-induced plant signals be transmitted aerially and through the rhizosphere? Biochemical Systematics and Ecology 29: Dugravot S, Brunissen L, Létocart E, Tjallingii WF, Vincent C et al. (2007) Local and systemic responses induced by aphids in

8 8 Ameline et al. Solanum tuberosum plants. Entomologia Experimentalis et Applicata in press. Eigenbrode SD, Ding H, Shiel PJ & Berger PH (2002) Volatiles released from Potato leafroll virus-infected potatoes are attractants and arrestants for its vector, Myzus persicae. Proceedings of the Royal Society of London. Series B, Biological Sciences 269: Gianoli E & Niemeyer HM (1997) Characteristics of hydroxamic acid induction in wheat triggered by aphid infestation. Journal of Chemical Ecology 23: Gonzales WL, Ramirez CC, Olea N & Niemeyer HM (2002) Host plant changes produced by the aphid Sipha flava: consequences for aphid feeding behaviour and growth. Entomologia Experimentalis et Applicata 103: Hardie J, Visser JH & Piron PGM (1994) Perception of volatiles associated with sex and food by different forms of the black bean aphid, Aphis fabae. Physiological Entomology 19: Harris K (1977) An ingestion-egestion hypothesis of noncirculative virus transmission. Aphids as Virus Vectors (ed. by K Harris & K Maramorosch), pp Academic Press, New-York, NY, USA. Jimenez-Martinez ES, Bosque-Perez NA, Berger PH & Zemetrar S (2004) Life history of the bird cherry-oat aphid, Rhopalosiphum padi (Homoptera: Aphididae), on transgenic and untransformed wheat challenged with barley yellow dwarf virus. Journal of Economic Entomology 97: Karban R & Baldwin I (1997) Induced Responses to Herbivory. University of Chicago Press, Chicago, IL,USA. Kay R (1976) Behavioural components of pheromonal aggregation in Aphis fabae Scopoli. Physiological Entomology 1: Kessler A & Baldwin I (2001) Defensive function of herbivoreinduced plant volatile emissions in nature. Science 291: Klingler J, Creasy R, Gao L, Nair R, Calix A et al. (2005) Aphid resistance in Medicago truncatula involves antixenosis and phloem-specific, inducible antibiosis, and maps to a single locus flanked by NBS-LRR resistance gene analogs. Plant Physiology 137: Kruzmane D, Jankevica L & Ievinsh G (2002) Effect of regurgitant from Leptinotarsa decemlineata on wound responses in Solanum tuberosum and Phaseolus vulgaris. Physiologia Plantarum 115: Macias W & Mink G (1969) Preference of green peach aphids for virus-infected sugarbeet leaves. Journal of Economic Entomology 62: McLean D & Kinsey J (1967) Probing behavior of pea aphid Acyrtosiphon pisum. I. Definitive correlation of electronically recorded waveforms with aphid probing activities. Annals of the Entomological Society of America 60: Nottingham SF, Hardie J, Dawson GW, Hick AJ, Pickett JA et al. (1991) Behavioural and electrophysiological responses of aphids to host and non-host plant volatiles. Journal of Chemical Ecology 17: Paré PW & Tumlinson JH (1999) Plant volatiles as a defense against insect herbivores. Plant Physiology 121: Petersen MK & Sandström JP (2001) Outcome of indirect competition between two aphid species mediated by responses in their common host plant. Functional Ecology 15: Pettersson J (1973) Olfactory reactions of Brevicoryne brassicae (L.) (Hom. Aph.). Swedish Journal of Agricultural Research 3: Pettersson J, Karunaratne S, Ahmed E & Kumar V (1998) The cowpea aphid Aphis craccivora, host plant odours and pheromones. Entomologia Experimentalis et Applicata 88: Pettersson J, Quiroz A, Stephansson D & Niemeyer HM (1995) Odour communication of Rhopalosiphum padi (L.) (Hom. Aph.) on grasses. Entomologia Experimentalis et Applicata 76: Pickett JA, Wadhams LJ, Woodcock CM & Hardie J (1992) The chemical ecology of aphids. Annual Review of Entomology 37: Pospisil J (1972) Olfactory orientation of certain phytophagous insects in Cuba. Acta Entomologica Bohemoslovia 69: Powell G, Tosh C & Hardie J (2006) Host plant selection by aphids: behavioral, evolutionary, and applied perspectives. Annual Review of Entomology 51: Prado E & Tjallingii WF (1994) Aphid activities during sieve element punctures. Entomologia Experimentalis et Applicata 72: Prado E & Tjallingii WF (1997) Effects of previous plant infestation on sieve element acceptance by two aphids. Entomologia Experimentalis et Applicata 82: Prado E & Tjallingii WF (1999) Effects of experimental stress factors on probing behaviour by aphids. Entomologia Experimentalis et Applicata 90: Quiroz A, Pettersson J, Pickett JA, Wadhams LJ & Niemeyer HM (1997) Semiochemicals mediating spacing behavior of bird cherry-oat aphid, Rhopalosiphum padi feeding on cereals. Journal of Chemical Ecology 23: Qureshi JA & Michaud JP (2005) Interactions among three species of cereal aphids simultaneously infesting wheat. Journal of Insect Science 5: 1 8. Sauge MH, Lacroze JP, Poëssel JL, Pascal T & Kervella J (2002) Induced resistance by Myzus persicae in the peach cultivar Rubira. Entomologia Experimentalis et Applicata 102: Tjallingii WF (1978) Electronic recording of penetration behaviour by aphids. Entomologia Experimentalis et Applicata 24: Tjallingii WF (1988) Electrical recording of stylet penetration activities. Aphids, Their Biology, Natural Enemies and Control, Vol. 2B (ed. by A Minks & P Harrewijn), pp Elsevier, Amsterdam, The Netherlands. Tjallingii WF & Hogen Esch T (1993) Fine structure of aphid stylet routes in plant tissues in correlation with EPG signals. Physiological Entomology 18: Tosh CR, Powell G & Hardie J (2002) Maternal reproductive decisions are independent of feeding in the black bean aphid, Aphis fabae. Journal of Insect Physiology 48: Voelckel C, Weisser WW & Baldwin IT (2004) An analysis of plant-aphid interactions by different microarray hybridization strategies. Molecular Ecology 13:

9 Immediate effects of plant damage on aphid behaviour 9 Walling LL (2000) The myriad plant responses to herbivores. Journal of Plant Growth Regulation 19: Will T & van Bel AJE (2006) Physical and chemical interactions between aphids and plants. Journal of Experimental Botany 57: Wool D & Hales DF (1996) Previous infestation affects recolonization of cotton by Aphis gossypii: induced resistance or plant damage? Phytoparasitica 24: Zhu-Salzman K, Salzman RA, Ahn J-E & Koiwa H (2004) Transcriptional regulation of Sorghum defense determinants against a phloem-feeding aphid. Plant Physiology 134: