Factors influencing the effectiveness of an attracticide formulation against the Oriental fruit moth, Grapholita molesta

Transcription

1 Blackwell Publishing, Ltd. Factors influencing the effectiveness of an attracticide formulation against the Oriental fruit moth, Grapholita molesta Maya L. Evenden 1, * & John R. McLaughlin 2 1 Department of Biology, West Chester University, West Chester, PA, USA; 2 IPM Tech Inc., 840 Main Campus Dr., 3590 Raleigh, NC 27606, USA Accepted: 28 April 2004 Key words: pheromone, attract and kill, lure and kill, orchard pest management, mating disruption, Lepidoptera, Tortricidae Abstract An attracticide formulation, LastCall OFM, was tested against the Oriental fruit moth Grapholita molesta (Busck) (Lepidoptera: Tortricidae) in replicated small plot field trials in apple, Malus domestica (Borkhausen), orchards in South-eastern Pennsylvania, USA. Attracticide treatments were applied using a calibrated hand pump, and treated plots were compared to similar untreated plots. Male moth activity was monitored using virgin female-baited traps, and the potential for reduction in mating activity was assessed using sentinel virgin females. A comparison of application rates showed that 1500 droplets per ha of the attracticide formulation was as effective as 3000 droplets per ha, and both application rates reduced captures in synthetic pheromone-baited traps for prolonged periods. Droplets placed either at high or low positions within the canopy significantly reduced trap capture and mating with sentinel females. In addition, the only sentinel females that mated in the treated plots were located in the untreated portion of the tree canopy. Mate finding behaviour was equally disrupted by formulations with and without insecticide. Therefore, under the test conditions, the mechanism by which the attracticide formulation worked was by disruption of male orientation, and not by the removal of males due to insecticide poisoning. Two field cage experiments tested the impact of population density on the competitiveness of the attracticide formulation compared to virgin females. A significant proportion of males were captured in female-baited traps at the highest female-to-droplet ratio tested. Equal proportions of males were captured in attracticide-baited traps at male moth densities of 10, 20, 40, and 80 males per cage. These results clarify some of the factors influencing the effectiveness and possible mechanisms of an attracticide management tactic against the Oriental fruit moth. Introduction *Correspondence: Maya Evenden, Department of Biological Sciences, CW 405 Biological Sciences Bldg., University of Alberta, Edmonton, AB, Canada T6G 2E9. Tel.: ; Fax: ; mevenden@ualberta.ca The potential for manipulating the behaviour of insect pests with pheromones to achieve economically acceptable control has been recognized for many years. Mating disruption by the release of a synthetic sex pheromone into the atmosphere has been investigated against many lepidopteran pests in different cropping systems (Cardé & Minks, 1995). Lepidopteran attracticides represent another pheromonebased technology which has been developed to provide better control at high population densities using reduced pheromone release rates, as compared to mating disruption formulations (Conlee & Staten, 1981). To date, lepidopteran attracticides have consisted of a combination of low concentrations of synthetic sex pheromone and pyrethroid insecticides that achieve a rapid knockdown effect (Butler & Las, 1983; Haynes et al., 1986; Miller et al., 1990; Downham et al., 1995; Charmillot et al., 1996; Charmillot & Hofer, 1997; Brockerhoff & Suckling, 1999; Suckling & Brockerhoff, 1999; Charmillot et al., 2000; Krupke et al., 2002). The optimal effectiveness of an attracticide depends on the exposure of the insect to the insecticide through source contact with the formulation (Charmillot et al., 1996; Suckling 2004 The Netherlands Entomological Society Entomologia Experimentalis et Applicata 112: 89 97,

2 90 Evenden & McLaughlin & Brockerhoff, 1999). This mode of action requires that the formulations are highly attractive, and that male moths follow synthetic false trails all the way to the source. Miller et al. (1986) demonstrated that male pink bollworm moths, Pectinophora gossypiella (Sanders) (Lepidoptera: Gelechiidae), followed false trails to pheromone-emitting fibres, as evidenced by the presence of wing scales on sticky fibres. An attracticide formulation with insecticide incorporated into the sticker proved more robust than pheromone alone against the pink bollworm (Conlee & Staten, 1981). In studies that have directly compared the disruption of male moth orientation with attracticide formulations both with and without insecticide, about 50% of the orientation disruption was the result of pheromone alone, and the additional 50% was from the removal of males from the population due to insecticide exposure (Charmillot et al., 1996; Suckling & Brockerhoff, 1999). The Oriental fruit moth Grapholita molesta (Busck) (Lepidoptera: Tortricidae), is a key pest of many peach and apple agroecosystems world-wide (Rothschild & Vickers, 1991). There is some evidence that the Oriental fruit moth would be a good candidate for control by an attracticide formulation. In small plot field trials, mating disruption of the Oriental fruit moth occurred at a lower release rate with the most attractive pheromone blend (Charlton & Cardé, 1981), suggesting that false-trail following to synthetic pheromone sources may be an important mechanism of orientation disruption for this species. Later wind tunnel studies showed that male Oriental fruit moths followed synthetic false trails successfully to formulations of the complete pheromone blend at release rates similar to, or slightly higher than, calling virgin females (Sanders & Lucuik, 1996; Valeur & Löfstedt, 1996). A recent attracticide formulation consists of a viscous paste that incorporates the insecticide, and an attractant, in a UV sensitive carrier material (Hofer & Brassel, 1992). This formulation has been registered for use against both the Oriental fruit moth and another tree fruit pest, the codling moth, Cydia pomonella (L.) (Lepidoptera: Tortricidae), in the USA, under the trade names LastCall OFM and LastCall CM, respectively. The success of the formulation in field trials against codling moth (Charmillot et al., 2000; Krupke et al., 2002), and preliminary lab and field experimentation against the light brown apple moth, Epiphyas postvittana (Walker) (Lepidoptera: Tortricidae), in New Zealand (Brockerhoff & Suckling, 1999; Suckling & Brockerhoff, 1999) suggests that it is a technology that could be effective in controlling the Oriental fruit moth. Our initial research (Evenden & McLaughlin, 2004) demonstrated that individual droplets of the registered attracticide formulation attract wild and laboratory-reared moths and expose them to insecticide via source contact. Here we have tested the attracticide in field settings to determine its most effective application rate and position, the impact of population density on the efficacy of the formulation, and the mechanism(s) invoked by the formulation. Materials and methods Attracticide formulations LastCall OFM formulations were prepared by D. Czokajlo (IPM Tech Inc., Portland, OR, USA). These consisted of a clear viscous paste comprised mainly of inert ingredients (93.8%). Oriental fruit moth pheromone incorporated into the formulations at a rate of 0.16% was a three-component blend consisting of 87.3% (Z)-8-dodecenyl acetate, 3.6% (E)-8 dodecenyl acetate, and 9.1% (Z)-8-dodecenyl alcohol. In the formulations with insecticide, permethrin was added at 6.0%. Formulations were dispensed as 50 µl droplets ( 50 mg) using a calibrated hand pump. Small plot experiments During the 2003 field season (April August), small plot experiments were established, following a randomized block design, in commercial apple orchards in South-eastern Pennsylvania within a 33 km radius of West Chester, PA (39 58 N, W). Three 0.1 ha ( m) plots, separated by a minimum of 40 m, were established in each of three experimental orchards. These orchards were used for Experiments 1 and 2. Different experimental sites were used in both Experiments 3 and 4, and in each case were located within separate, individual orchards. Each site was separated by 100 m, and plots within sites were separated by 40 m. Individual treatments were randomly assigned to plots in each of the three sites per experiment. Treatments were applied to the central leader and branches of apple trees. Treatments were assessed with sentinel virgin females, and virgin female-baited traps. Female moths were obtained from a laboratory colony maintained on a lima bean-based diet at L16:D8 and 24 C. Pupae were separated by sex, and individual female pupae were held in 30 ml cups and monitored daily. Moths were provided with a water source until use at 1 5 days post-eclosion. Sentinel virgin females were placed individually in small mating cages made from cone-orifice traps (6 cm tall by 2.5 cm diameter). Cages were modified to provide the moth with a water source (Fitzpatrick & Troubridge, 1993), and glued to the surface of a Petri dish lid (10 cm diameter) that was fitted with a wire hanger for suspension within a tree. Virgin femalebaited traps consisted of Intercept A traps (IPM Tech Inc.) fitted with a removable sticky liner, and baited with a small mesh bag (9 6.5 cm) containing one virgin female moth. Both mating cages and mesh bags containing virgin females were transported to field sites in a refrigerated container.

3 Oriental fruit moth attracticide 91 Six sentinel virgin females and six virgin female-baited traps were suspended within the tree canopy at the plot centre during each of three, 4-day assessment periods per experiment. Three sentinel females and three female-baited traps were positioned at both low (head height) and high (upper third of the canopy) positions within the canopy. The placement of each female and trap was marked with flagging tape and maintained in subsequent assessments. Females were distributed evenly among treatments by age. The mating status of sentinel females recovered from the experimental plots was determined by dissection to reveal the presence (mated) or absence (virgin) of a spermatophore in the bursa copulatrix. Virgin male moths were released in equal numbers in each plot during each assessment period (except in Experiment 1) to augment the natural populations. Male moths were obtained from the laboratory colony and housed in the same manner as outlined for females until use, 1 6 days post-eclosion. Males were chilled at 5 C for 30 min and then transferred in groups of to Petri dishes (5 cm in diameter) that were fitted with a wire hanger for suspension within the tree canopy. Petri dishes containing male moths were transported to field sites in refrigerated containers. The lids of the Petri dishes were removed and two Petri dishes were suspended within the mid-canopy in the centre row of each plot, 7 m on either side of the plot centre. Males were distributed evenly among treatments by age. Experiment 1 compared plots treated with different rates of attracticide formulation to untreated control plots. Treatments were applied at either 1500 or 3000 droplets per ha. Droplets were applied in the low to mid-canopy on the centre leader and branches of the trees. The three assessment periods were taken days post-treatment. During the first assessment period no laboratory-reared virgin males were released in the treated plots. Due to low male capture and mating rates of sentinel females in the first assessment period, we decided to supplement the populations during the second and third assessment periods. In each of the second and third assessment periods, 30 virgin males were released in each plot at the time of female placement within the plots. During the third assessment period, plots in only one site were assessed due to a shortage of reared moths at that time for a total n = 7. The proportions of sentinel females that mated were arcsine-square root transformed, and the number of males captured in virgin female-baited traps were log (x + 1) transformed to satisfy requirements for normality and homoscedasticity (Zar, 1984). Both data sets were then analyzed using a randomized block design ANOVA with orchard specified as a random variable and assessment period treated as a repeated measure (PROC MIXED, SAS, 1996). Analysis of variance was followed by least square means tests (SAS, 1996) to compare individual treatments. Experiment 2 compared the capture of male Oriental fruit moths over a 16-week period in synthetic pheromonebaited traps to determine the longevity of the attracticide formulations assessed in Experiment 1. One Intercept A trap was baited at field sites with a red rubber septum monitoring lure containing mg of the same Oriental fruit moth pheromone blend used in the attracticide formulations (IPM Tech Inc.). The traps were placed 1.5 m off the ground in the centre tree of each plot immediately following attracticide treatment of the plot. Trap captures were assessed at weekly intervals, but traps were not present in plots during treatment assessment with sentinel virgin females and virgin female-baited traps (Experiment 1). Moths were removed from sticky liners weekly, and the liners were replaced as needed. Pheromone lures were replaced at 4-weekly intervals. Male moth catches in the synthetic-baited traps were log(x + 1) transformed and compared using a two-way ANOVA (assessment period and treatment) with orchard specified as a random factor and assessment period treated as a repeated measure (PROC MIXED, SAS, 1996). Analysis of variance was followed by least square means tests (SAS, 1996) to compare individual treatment by assessment period interactions. Experiment 3 tested the hypothesis that droplet position influences the effectiveness of the attracticide. For this experiment, sites within the same orchard with similar standard tree plantings (trees 4 m high) were chosen so there was a consistent difference between the high and low droplet positions. Treatments consisted of attracticide applied at 1500 droplets per ha at low (head height) and high (upper third of canopy) positions, and a non-treated control. The three assessment periods occurred 1 20 days post-treatment. Forty-five male moths were released in each plot during each of the three assessment periods. All three sites were assessed during each period for a total of n = 9. Data sets were transformed as in Experiment 1 and analyzed using a 2-way ANOVA (treatment and female position) with site specified as a random factor and assessment period treated as a repeated measure (PROC MIXED, SAS, 1996). Analysis of variance was followed by least square means tests (SAS, 1996) to compare individual treatments. Experiment 4 tested the hypothesis that attracticide treatment results in the disruption of mate finding and mating by removal of males from the population through permethrin exposure. Attracticide formulations with and without the 6% permethrin component were applied to plots at a treatment rate of 3000 droplets per ha, with droplets applied at both low (2/3 of droplets) and high (1/3 of droplets) positions within the canopy and compared to untreated control plots. The three assessment periods

4 92 Evenden & McLaughlin occurred 3 15 days post-treatment. Twenty, 35, and 30 males were released in each plot during each assessment period, respectively. All three sites were assessed during each period, for a total of n = 9. The effect of treatments on the proportion of sentinel females that mated and the number of males captured in female-baited traps were analyzed as in Experiment 1. Field cage experiments Two field cage experiments were conducted to determine the impact of population density on the number of individuals attracted to droplets of attracticide. Four field cages ( m) were erected >25 m apart on grassy locations on the West Chester University campus (39 58 N W). The cages were placed close to buildings so that there was no direct airflow between them. Within each cage, seven 1.9-l buckets fitted with lids and filled with water were placed in an H configuration, with each bucket separated by 60 cm. The lid of each bucket was punctured, and freshly cut apple branches were placed in each bucket to create a small ( 60 cm-high) canopy. Branches remained in the cages for one replicate, and were replaced at the beginning of each new replicate in both experiments. Each experiment consisted of four, one-night replicates. Treatments were randomly assigned to the cages and were re-randomized over time following a latin square randomization, so that each treatment occupied each cage at one time. During each replicate, 1 5-day-old virgin male moths were released at the cage centre from a 5 cm diameter Petri dish placed within the branch canopy. Males were released in the late afternoon and given 1 h to acclimate to the cage conditions. Males were distributed evenly among treatments by age. Males were reared and handled as outlined for the field experiments, except that they were marked with one of four colours of Day-Glo UV fluorescent powder (Switzer, Cleveland, OH, USA) to ensure individuals were enumerated in only one replicate. Traps were introduced to the cages 1 h after the males had been released, and remained in the cages overnight. Attracticide-baited traps consisted of Intercept A traps fitted with a sticky liner and baited with a 50 µl droplet weighing between 42.5 and 52.5 mg, positioned in the centre of a 1 2 cm piece of aluminium foil and suspended vertically by a short length of wire from the top of the trap. Virgin female-baited traps consisted of the same trap type, but baited with an individual 2 4-dayold virgin female in a mesh bag suspended from the top of the trap. Females were reared and handled as outlined in the field experiments. Sticky traps were positioned on bamboo stakes placed adjacent to buckets within the canopy, 60 cm off the ground and separated by 60 cm. The attracticide-baited trap was always positioned in the centre of the trap array, and the trap locations were maintained between replicates in both experiments. Traps were removed from cages the following morning, and males captured in each trap were counted and recorded. Males that were not captured were removed from the cage with a vacuum. Experiment 5 tested the hypothesis that the density of calling virgin females influences the proportion of males within a population that orient to attracticide droplets. During each replicate, 25 virgin male moths were released within the canopy at the centre of the cage. Treatments consisted of: (1) one attracticide-baited trap (control); (2) one attracticide-baited trap and two virgin-female baited traps; (3) one attracticide-baited trap and four virgin femalebaited traps; and (4) one attracticide-baited trap and eight virgin female-baited traps. Experiment 6 tested the hypothesis that the density of virgin male moths influences the proportion of males attracted to attracticide droplets. During each replicate, one attracticide-baited trap and eight virgin female-baited traps were positioned in each cage. The four treatments consisted of the release of 10, 20, 40, and 80 marked, virgin males. Field cage data were collected as the proportion of male moths that responded within each replicate. Proportions were arcsine-square root transformed (Zar, 1984) and treatments were compared using a latin square design ANOVA (PROC GLM, SAS, 1996). Analysis of variance was followed by least square means tests (SAS, 1996) to compare individual treatments. Results Small plot experiments In Experiment 1, a total of 88 males were captured in female-baited traps, and a total of 21 of 114 sentinel virgin females were mated across all treatments. The treatment of small plots with attracticide at rates of 1500 and 3000 droplets per ha significantly and equally reduced the number of males captured in virgin female-baited traps and the proportion of sentinel females that mated as compared to untreated control plots (Figure 1A,B). In Experiment 2, these same formulations shut down trap capture in synthetic pheromone-baited traps for 7 weeks post-treatment. However, due to the low population pressure at our sites, captures in control plots were only significantly greater than both treated plots for the first (P<0.0001) and second (P = ) weeks post-treatment. The capture of males in the 1500 droplet per ha treatment was also significantly lower than in control plots at 9 weeks (P = ) and 14 weeks (P = ) post-treatment. In Experiment 3, a total of 277 males were captured in female-baited traps, and 12 of the 134 sentinel virgin females mated across all treatments. Treatment with attracticide at 1500 droplets per ha positioned either high or low in the

5 Oriental fruit moth attracticide 93 throughout the canopy significantly and equally reduced the number of males captured in female-baited traps, and the proportion of females that mated (Figure 3A,B). Although no sentinel females mated in the plots treated with the formulation without insecticide, this was not significantly different from the proportion of females that mated in plots treated with formulations containing insecticide which represents two mated females out of 43 tested. Field cage experiments The attracticide-baited traps in Experiment 5 captured the majority of recovered moths at all of the attracticide-tovirgin female ratios tested. However, when eight females were present in the cages, the proportion of responders captured in the attracticide-baited trap was significantly reduced (Figure 4A), demonstrating that a significant proportion of the responders were captured in female-baited traps. In Experiment 6, the attracticide-baited traps captured a similar proportion of responding males with a constant female density of eight per cage, regardless of the male moth density within each cage (Figure 4B). Figure 1 (A) Mean (+ SE) number of males captured per femalebaited trap; (B) mean (+ SE) proportion of sentinel females mated in plots treated with attracticide; n = 7. Bar indicated with * is significantly different, least means squares (P<0.05). canopy significantly reduced the number of males captured in virgin female-baited traps as compared to non-treated control plots (P<0.0001). However, trap capture was not influenced by the position of the female-baited trap within the canopy (P = ), and there was no treatment-bytrap position interaction effect (P = ) (Figure 2A). The proportion of females that mated was significantly greater in the control plots than in either of the treated plots (P = ). All of the females that mated in plots treated with droplets in the low position were located high in the canopy. Similarly, all of the females that mated in plots treated with droplets at the high position were located low in the canopy. This resulted in a significant treatmentby-female position interaction effect (P = ) on the proportion of sentinel females that mated (Figure 2B). In Experiment 4, a total of 287 males were captured in female-baited traps and 15 of the 133 sentinel virgin females mated across all treatments. Mate-finding behaviour was equally disrupted by formulations with and without insecticide. The treatment of small plots with formulations with and without permethrin at 3000 droplets per ha positioned Figure 2 (A) Mean (+ SE) number of males captured per femalebaited trap; (B) mean (+ SE) proportion of sentinel females mated in plots treated with attracticide droplets at high and low positions within the tree canopy; n = 9.

6 94 Evenden & McLaughlin release rate from the formulation may be less crucial than its point source density. The male moths ability to follow synthetic false trails to the source should be most effective when the formulation point source density is low, providing discrete plumes and a low chance of camouflage and neurophysiological effects caused by uniformly dispersed pheromone. In 0.08 ha plots treated with an attracticide formulation against the pink bollworm, males contacted point sources more often at densities of 112, 218, and 450 sources per plot than at the higher density of 900 sources per plot (Miller et al., 1990). Similarly, the proportion of fibres that were contacted in a wind tunnel decreased with increasing density (Miller et al., 1990). However, the point source density comparisons of the pink bollworm attracticide differed from our Figure 3 (A) Mean (+ SE) number of males captured per femalebaited trap; (B) mean (+ SE) proportion of sentinel females mated in plots treated with attracticide with (LastCall) and without (OFM) insecticide; n = 9. Bar indicated with * is significantly different, least means squares (P<0.05). Discussion Because LastCall OFM is now registered and available for use in the USA, it is important that growers use it in the most effective and economical way. Experiment 1 revealed that, at low to moderate population densities, the recommended application rate could be reduced, as the half concentration rate (1500 droplets per ha) was as effective as the full rate (3000 droplets per ha) at reducing trap catch in female-baited traps and mating of sentinel females. Disruption of male Oriental fruit moth orientation to calling virgin females has been shown to be dependent on the concentration of synthetic sources in both wind tunnel (Sanders & Lucuik, 1996; Valeur & Löfstedt, 1996) and field experiments (Charlton & Cardé, 1981; Rothschild & Vickers, 1991; and references therein). However, attracticide formulations use much less pheromone than mating disruption formulations, and their optimal effectiveness depends on insect exposure to insecticide through source contact with the formulation (Charmillot et al., 1996; Suckling & Brockerhoff, 1999). Therefore, overall pheromone Figure 4 (A) Mean (+ SE) proportion of total Oriental fruit moth males captured in field cages by attracticide-baited traps at varying female densities; n = 4. Bars with the same letter are not significantly different, least means squares (P>0.05). (B) Mean (+ SE) proportion of total Oriental fruit moth males captured in field cages by attracticide-baited traps at varying male densities; n = 4. Bars with the same letter are not significantly different, least means squares (P>0.05).

7 Oriental fruit moth attracticide 95 experiments in that the pheromone release rate remained constant, despite varying the number of point sources (Miller et al., 1990). Our treatments did not discriminate between the effects of pheromone release rate and the number of point sources. In Experiment 1, both application rates may have been equally effective as a result of enhanced false trail following and source contact with droplets at the lower point source density. Increased source contact may have compensated for the lower point source density and pheromone release rate, and produced similar results to the higher application rate. However, the density of attracticide sources must also be high enough to compete with the natural pheromone plumes produced by calling females. In small plots treated with attracticide against the codling moth, interference with mate-finding behaviour was not observed at point source densities below 500 droplets per ha (Krupke et al., 2002). Likewise, a point source density of 450 droplets per ha of a similar droplet attracticide formulation against the light brown apple moth resulted in 88 96% reduction in male moth capture in synthetic pheromone-baited traps (Suckling & Brockerhoff, 1999). The plots treated with 1500 and 3000 droplets per ha were monitored for 16 weeks after treatment application in an attempt to determine the longevity of the formulation. However, low population densities and unseasonably inclement weather 3 8 weeks after treatment precluded us from obtaining relevant data during this period. For these reasons, male moth capture in plots treated with both application rates of attracticide was significantly different from captures in control plots for only 2 weeks post-treatment. The capture of males in the 1500 droplet per ha treatment was also significantly lower than in the control plots at 9 and 14 weeks post-treatment. Our results, although incomplete, agree with similar season-long monitoring of orchards treated with attracticide against codling moth. No male codling moths were captured in attracticide-treated plots for 48 days following application, and catches remained low for the remainder of the season (Charmillot et al., 2000). Experiment 3 revealed an interesting effect of droplet placement within the orchard canopy. Droplets placed at either high or low positions within the canopy significantly reduced trap capture and mating with sentinel females (Figure 2A,B). Male Oriental fruit moth capture in synthetic pheromone-baited traps was higher in traps positioned at 2 m than 1 m and 4 m off the ground, regardless of tree height (Rothschild & Minks, 1977), suggesting that most flight activity occurs in the mid- to upper-canopy. Field observations conducted in peach orchards suggested that most Oriental fruit moth mating activity occurs within the upper canopy (Rothschild & Vickers, 1991). In our study, all of the sentinel females that mated in the treated plots were located in the untreated portion of the canopy (Figure 2B). Similarly, the placement of codling moth mating disruption dispensers low in the canopy resulted in a greater proportion of sentinel females that mated high in the canopy (Weissling & Knight, 1995). However, the proportion of codling moth females that mated high in the canopy was also greater in control plots, and pheromone dispensers positioned high in the canopy reduced the mating of females throughout the canopy (Weissling & Knight, 1995). To ensure disruption of the mate-finding behaviour of Oriental fruit moths throughout the canopy, an alternating attracticide droplet position is recommended. Comparison of attracticide formulations with and without permethrin applied at the recommended application rate of 3000 droplets per ha throughout the canopy revealed that mate-finding and mating behaviour were equally disrupted by both formulations, regardless of insecticide content. Therefore, it appears that the main mechanism by which this formulation works in small plots with low to moderate population densities is by the disruption of male moth orientation, and not by removal of males due to insecticide poisoning. These results contradict the findings of similar experiments on other tree fruit tortricids (Charmillot et al., 1996; Suckling & Brockerhoff, 1999). A comparison of the trap catch of male light brown apple moths in plots treated with attracticide vs. caged attracticide, in which males are prohibited from touching the insecticideladen droplets, showed that about 50% of the effect of trap suppression was caused by disruption of male moth orientation, and 50% was due to the removal of males from the population due to insecticide exposure (Suckling & Brockerhoff, 1999). However, the attracticide droplets in the light brown apple moth study were applied at a lower density (450 droplets per ha) than in our study, and might have enhanced false trail following to the point source and subsequent exposure to insecticide. Because orientation to pheromone sources by male Oriental fruit moths is strictly defined by a lower and upper pheromone dose threshold (Baker & Cardé, 1979; Baker et al., 1981; Kuenen & Baker, 1982) males may exhibit neurophysiological mechanisms of disruption (Sanders & Lucuik, 1996) even at low point source densities of attracticide. Mating of tethered virgin female Egyptian cotton leafworm, Spodoptera littoralis (Boisduval) (Lepidoptera: Noctuidae), was equally disrupted by attracticide formulations with and without insecticide applied to cotton plants at the low density of 500 point sources per ha (Downham et al., 1995). At low population densities, the pink bollworm was equally disrupted by formulations with and without insecticide. However, the insecticide-laden formulation had a better efficacy at higher population densities (Conlee & Staten, 1981). Therefore,

8 96 Evenden & McLaughlin just as is the case for pheromone-based mating disruption (Cardé & Minks, 1995), the mechanisms of control by attracticide formulations will vary among pest species, cropping systems, and population densities. The effectiveness of an attracticide as a control agent will depend on the competition provided by natural sources of pheromone, and might be expected to decrease as the population density increases (Charmillot et al., 2000; Krupke et al., 2002). In Experiment 5, we demonstrated that attracticide droplets were competitive with calling females at ratios of 1 : 2 and 1 : 4 in field cages. However, at a dropletto-female ratio of 1 : 8, there was a significant reduction in the proportion of captured males in the droplet-baited trap (Figure 4A), emphasizing the importance of point source competition with naturally occurring pheromone plumes. Similar effects of female population density were observed in field cage studies of an attracticide formulation against the codling moth (Krupke et al., 2002). Interestingly, the single attracticide-baited trap placed within an array of eight female-baited traps in Experiment 6, captured the same proportion of males, regardless of the male population density (Figure 4B). Increased pink bollworm male density in a wind tunnel bioassay revealed a decrease in mortality from exposure to attracticide (Miller et al., 1990). The decreased mortality was thought to be due to several factors, including increased competition at the attracticide source resulting in less time spent on the insecticide laden source and a build up of moth scales preventing contact with the insecticide (Miller et al., 1990). Our bioassay simply measured attraction to droplet- vs. female-baited traps, and males did not experience competition at the droplet or a heavily deteriorated droplet surface. However, because an equal proportion of the captured males flew to female-baited traps at each male population density tested, more males flew to females at high population densities, and this would impact the effectiveness of an attracticide in field applications. These small plot field trials and field cage experiments have demonstrated several important factors that could influence the effectiveness of the commercial application of an attracticide formulation against the Oriental fruit moth. At low to moderate population densities, a reduction in the recommended application rate is unlikely to alter the efficacy of this formulation, and may enhance the insecticide exposure of males through increased false trail following to droplets. Droplets need to be distributed throughout the tree canopy for complete disruption of mate-finding behaviour. The attracticide formulation will also be most effective at low pest population densities. Further work is needed to determine whether suppression of mating will result in reduced larval populations and fruit damage in a commercial setting. Acknowledgements We thank D. Czokajlo (IPM Tech Inc.) for formulating the LastCall OFM treatments used in this study. IPM Tech Inc. provided traps for use in all field experiments. P.W. Shearer (Rutgers, The State University of New Jersey) provided us with initial insect material to start our laboratory colony. Orchardists in the Chester/Delaware Counties (Pennsylvania, USA) Fruit Growers Association permitted use of their orchards for field experiments. R. French, M. Kline, and A. Seiden assisted us with insect rearing and field work. Two anonymous reviewers made helpful suggestions on an earlier draft of this manuscript. This work was supported by a US Department of Agriculture NE Sustainable Agriculture Research and Education Partnership Grant to M.L.E., a Pennsylvania State System of Higher Education Faculty Professional Development Council Grant to M.L.E., and a West Chester University Presidential Initiative Funding Grant to M.L.E. References Baker TC & Cardé RT (1979) Analysis of pheromone-mediated behavior in male Grapholitha molesta, the Oriental fruit moth (Lepidoptera: Tortricidae). Environmental Entomology 8: Baker TC, Meyer W & Roelofs WL (1981) Sex pheromone dosage and blend specificity of response by Oriental fruit moth males. Entomologia Experimentalis et Applicata 30: Brockerhoff EG & Suckling DM (1999) Development of an attracticide against light brown apple moth (Lepidoptera: Tortricidae). Journal of Economic Entomology 92: Butler GD & Las AS (1983) Predaceous insects: effects of adding permethrin to the sticker used in gossyplure applications. Journal of Economic Entomology 76: Cardé RT & Minks AK (1995) Control of moth pests by mating disruption: successes and constraints. Annual Review of Entomology 40: Charlton RE & Cardé RT (1981) Comparing the effectiveness of sexual communication disruption in the Oriental fruit moth (Grapholitha molesta) using different combinations and dosages of its pheromone blend. Journal of Chemical Ecology 7: Charmillot PJ & Hofer D (1997) Control of codling moth, Cydia pomonella L., by an attract and kill formulation. Technology transfer in mating disruption. IOBC WPRS Bulletin 20 (1): Charmillot PJ, Hofer D & Pasquier D (2000) Attract and kill: a new method for control of the codling moth Cydia pomonella. Entomologia Experimentalis et Applicata 94: Charmillot PJ, Pasquier D, Scalco A & Hofer D (1996) Essais de lutte contre le carpocapse Cydia pomonella L. par un procédé attracticide. Mitteilungen der Schweizerischen Entomologischen Gesellschaft 69: Conlee JK & Staten RT (1981) Device for Insect Control. US Patent 4,671,010.

9 Oriental fruit moth attracticide 97 Downham MCA, McVeigh LJ & Moawad GM (1995) Field investigation of an attracticide control technique using the sex pheromone of the Egyptian cotton leafworm, Spodoptera littoralis (Lepidoptera: Noctuidae). Bulletin of Entomological Research 85: Evenden ML & McLaughlin JR (2004) Initial development of an attracticide formulation against the Oriental fruit moth Grapholita molesta (Lepidoptera: Tortricidae). Environmental Entomology 33: Fitzpatrick SM & Troubridge JT (1993) A modified live-trap for assessing mating disruption of microlepidoptera. Canadian Entomologist 125: Haynes KF, Li WG & Baker TC (1986) Control of pink bollworm moth (Lepidoptera: Gelechiidae) with insecticides and pheromones (attracticide): lethal and sublethal effects. Journal of Economic Entomology 79: Hofer D & Brassel J (1992) Attract and kill to control Cydia pomonella and Pectinophora gossypiella. Bulletin of the OILB/ SROP 15: Krupke CH, Roitberg BD & Judd GJR (2002) Field and laboratory responses of male codling moth (Lepidoptera: Tortricidae) to a pheromone-based attract-and-kill strategy. Environmental Entomology 31: Kuenen LPS & Baker TC (1982) Optomotor regulation of ground velocity in moths during flight to sex pheromone at different heights. Physiological Entomology 7: Miller E, Jones E & Staten RT (1986) The use of moth scales to determine male pink bollworm visitation to individual pheromone dispensers in a mating disruption system. Southwestern Entomologist 2: Miller E, Staten RT, Nowell C & Gourd J (1990) Pink bollworm (Lepidoptera: Gelechiidae): point source density and its relationship to efficacy in attracticide formulations of gossyplure. Journal of Economic Entomology 83: Rothschild GHL & Minks AK (1977) Some factors influencing the performance of pheromone traps for Oriental fruit moth in Australia. Entomologia Experimentalis et Applicata 22: Rothschild GHL & Vickers RA (1991) Biology, ecology and control of the oriental fruit moth. World Crop Pests, Vol. 5: Tortricid Pests, Their Biology, Natural Enemies and Control (ed. by W Helle, LPS van der Geest & HH Evenhuis), pp Elsevier, Amsterdam, The Netherlands. Sanders CJ & Lucuik GS (1996) Disruption of male Oriental fruit moth to calling females in a wind tunnel by different concentrations of synthetic pheromone. Journal of Chemical Ecology 22: SAS Institute (1996) SAS User s Guide: Basics, 6.03 edn. SAS Institute, Cary, NC, USA. Suckling DM & Brockerhoff EG (1999) Control of light brown apple moth (Lepidoptera: Tortricidae) using an attracticide. Journal of Economic Entomology 92: Valeur PG & Löfstedt C (1996) Behaviour of male oriental fruit moth, Grapholita molesta, in overlapping sex pheromone plumes in a wind tunnel. Entomologia Experimentalis et Applicata 79: Weissling TJ & Knight AL (1995) Vertical distribution of codling moth adults in pheromone-treated and untreated plots. Entomologia Experimentalis et Applicata 77: Zar JH (1984) Biostatistical Analysis, 2nd edn. Prentice Hall, Englewood Cliffs, USA.