Blackwell Publishing, Ltd.
Factors influencing the effectiveness of an attracticide formulation against the Oriental fruit moth, Grapholita molesta Maya L. Evenden1,* & John R. McLaughlin2 1
Department of Biology, West Chester University, West Chester, PA, USA; 2IPM 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 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 pheromone*Correspondence: Maya Evenden, Department of Biological Sciences, CW 405 Biological Sciences Bldg., University of Alberta, Edmonton, AB, Canada T6G 2E9. Tel.: +1 780 492 1873; Fax: +1 780 492 7150; E-mail:
[email protected]
based 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, 2004
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& 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, 75°38′W). Three 0.1 ha (33.3 × 33.3 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.
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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 10–25 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 16–53 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 0.111 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
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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 (1.83 × 1.83 × 1.83 m) were erected >25 m apart on grassy locations on the West Chester University campus (39°58′N 75°38′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
