Polyvinyl chloride polymer (PVC) dispensers loaded with ethyl (E,Z)-2,4-decadienoate (pear ester) plus the sex pheromone, (E,E)-8,10-dodecadien-1-ol (codlemone) of codling moth, Cydia pomonella (L.), were compared with PVC dispensers and a commercial dispenser (Isomate-C Plus) loaded with codlemone. Evaluations were conducted in replicated plots (0.1-0.2 ha) in apple, Malus domestica (Borkhausen) during both generations of codling moth from 2007 to 2009. Dispensers were applied at 1,000 ha−1. Male captures in traps baited with virgin female moths and codlemone lures were recorded. Residual analysis of field-aged dispensers over both moth generations was conducted. Dispensers exhibited linear declines in release rates of both attractants, and pear ester was released at a significantly higher rate than codlemone during both time periods. The proportion of virgin female-baited traps catching males was significantly lower with combo dispenser TRE24 (45/110, mg codlemone/mg pear ester) during the second generation in 2007 and the combo dispensers TRE144 (45/75) and TRE145 (75/45) during the first generation in 2008 compared with Isomate-C Plus. Similarly, male catches in female-baited traps in plots treated with the combo dispensers TRE144 during the first generation in 2008 and TRE23 (75/110) during the second generation, in 2007 were significantly lower than in plots treated with Isomate-C Plus. No significant differences were found for male catches in codlemone-baited traps in plots treated with Isomate-C Plus and any of the combo dispensers. However, male catches were significantly lower in plots treated with Cidetrak CM (codlemone-only dispenser) than the combo TRE144 dispenser during both generations in 2009.
Mating disruption with hand-applied dispensers has been widely adopted to manage codling moth, Cydia pomonella (L.) (Witzgall et al. 2008). Competitive attraction of males to synthetic dispensers versus native calling females is considered to be a primary behavioral mechanism of how hand-applied dispensers disrupt mating of codling moth (Miller et al. 2010). We have hypothesized (Light et al. 2001) that the observed increased attractive response of male codling moth to blends of (E,E)-8, 10-dodecadien-1-ol, codlemone, and pear ester, ethyl (E, Z)-2,4-decadienoate, could be used to improve the effectiveness of dispensers applied for mating disruption (Minks and Cardé 1988, Cardé and Minks 1995).
Several previous field studies have evaluated the use of hand-applied polyvinyl chloride (PVC) polymer dispensers loaded with high rates of a blend of codlemone and pear ester for mating disruption (Light and Knight 2005, Steliniski et al. 2007, Knight et al. 2011a, Bohnenblust et al. 2011). Unfortunately, treatments, experimental protocols, and results have all been variable. For example, initial studies conducted in California during 2004 found that combo dispensers loaded with either 264 or 537 mg of these co-attractants significantly reduced male catches in codlemone-baited traps and lowered fruit injury in small plots compared with the use of Isomate-C TT dispensers loaded with only codlemone (Light and Knight 2005). However, these results were confounded by the use of different dispenser densities, an important variable previously shown to affect the effectiveness of disruption (Epstein et al. 2006). Interestingly, the co-application of codlemone and pear ester in a single dispenser reduced the incidence of multiple-mated females relative to similar plots treated only with codlemone (Light and Knight 2005). This result may be important because multiple mating by female codling moth has been identified as a significant positive factor affecting fecundity (Knight 2007a). Encouraging results with combo dispensers were also reported in 2005 when dispensers loaded with 290 mg of codlemone and pear ester significantly reduced mating of tethered females over the initial 4-wk period compared with Isomate-C Plus dispensers loaded with only codlemone (Steliniski et al. 2007). However, no difference was found between the use of these dispensers in the mating success of females over the subsequent 3 wk of the test. This temporal pattern could be associated with changes that occur in the blend or ratio of constituents released by combo dispensers. For example, the combo dispensers tested in 2006 released pear ester initially at levels 10-fold higher than codlemone, but this ratio dropped continuously throughout the season, until both attractants were released at nearly the same rate after 18 wk (Knight et al. 2011a). Further trials conducted in Michigan in 2006 did not find any significant reductions in either the mating of tethered females or male catch in codlemone-baited traps with combo dispensers loaded with either a 170 or 340 mg blend compared with codlemone-only dispensers (Stelinski et al. 2007). In addition, plots treated with the higher-load combo dispenser in Washington State during 2006 had significantly lower male moth catch in codlemone-baited traps than similar plots treated with Checkmate CM 1000XL dispensers loaded only with codlemone (Knight et al. 2011a). Again, dispensers were tested at different application densities. Concurrently, a 50% reduction in male catch by virgin female-baited traps in plots treated with combo versus codlemone-only dispensers was not statistically significant, because of the low number of replicates and high variability that occurred among plots. Male catches in codlemone-baited traps were significantly lower in Michigan plots treated with the same density of a 340 mg combo dispenser versus Isomate-C Plus during the first generation in 2007 (Knight et al. 2011a). Male moth catches in codlemone-baited traps were also significantly lower in Pennsylvania apple plots treated with a 185 mg combo dispenser in 2007 and 2008 compared with plots treated with CM Disrupt Micro-Flakes loaded with only codlemone; but was not different from catches in plots treated with Isomate CM/OFM TT loaded with codlemone applied at half the rate (Bohnenblust et al. 2011).
This large group of studies with either positive or neutral results comparing the effectiveness of dispensers loaded with pear ester and codlemone versus codlemone alone were sufficiently interesting to stimulate further studies aimed to refine the use of combo dispensers for management of codling moth. Herein, data are presented from a series of replicated, field trials conducted from 2007 to 2009 in Washington State to evaluate the effect of different loadings and ratios of pear ester and codlemone in comparison to codlemone alone when tested at the same dispenser density (1,000 ha−1). Six formulations of combo PVC dispensers loaded with 90–155 mg of co-attractants, PVC dispensers loaded only with codlemone (75 or 120 mg), and the polyethylene dispenser Isomate-C Plus loaded only with codlemone (96.5 mg) were evaluated. Male catches by both virgin female-baited and synthetic lure-baited traps were compared between dispenser treatments and an untreated control. Residual analyses of field-aged PVC dispensers were conducted each year to estimate the release rates of attractants during each moth generation.
Materials and Methods
Studies were conducted with experimental and commercial dispensers. Experimental dispensers (TRE#, Trécé Inc., Adair, OK) were constructed of a proprietary, PVC polymer and loaded as blends of 90–185 mg of codlemone and pear ester or with only 75 mg of codlemone. The registered PVC dispenser, Cidetrak CM was formulated with 120 mg codlemone and was tested in 2008 and 2009. The effectiveness of these dispensers was compared in 2007 and 2008 with a polyethylene reservoir dispenser, Isomate-C Plus (Pacific Biocontrol, Vancouver, WA) formulated with 96.5 mg of codlemone.
Analysis of Dispensers.
PVC dispensers were placed each year in a ‘Fuji’ apple orchard located at the USDA Research Farm 13 miles east of Moxee, WA (46° 34′ N, 120° 04′ W) for residual analysis. Dispensers during 2007 and 2008 were attached to small branches on 25 May and 12 May, respectively. Dispensers during 2009 were attached to a specialized metal hanger (Trécé Inc.) that was hung on the tree. Dispensers (N=2) were collected every 2 wk and placed in sealed foil freezer pouches, stored at −18°C, and shipped to Trécé's analytical lab for extraction and testing. Dispensers collected on 6 July and 17 September 2007, 7 July and 15 September 2008, and 8 July and 11 September 2009 were analyzed for this study. TRE144 dispensers that were field-tested in 2009 were not analyzed. Isomate-C Plus dispensers were field-aged with similar methods in an apple orchard near Wenatchee, WA, in 2008 and in the same Moxee orchard as the PVC dispensers in 2009. These dispensers were collected on 12 May, 7 July, and 20 August in 2008; and 28 May, 8 July, and 16 September in 2009.
The dispenser extraction protocol was similar for both PVC and Isomate dispensers. PVC dispensers were ground to a homogeneous size (≈2 × 5 mm). A 1 g sample was extracted (1 h sonication) in 15 ml methanol with 20.0 mg of methyl tetradecanoate added as an internal standard. Isomate-C Plus dispensers were cut into 3–5 mm lengths and placed with methanol in 20 ml scintillation vials with 100 mg of methyl tetradecanoate. The cut dispenser tubes were washed four times with anhydrous methanol and the final washing was sonicated for 1 h. Two milliliters aliquots of extract for each dispenser were transferred to gas chromatography (GC) vials for analysis. Residual analysis was conducted with 1-μl injections into a 7890 GC (Agilent, Palo Alto, CA) equipped with a split/splitless injector, a flame ionization detector, and a RTX-5 (Restek, Bellefonte, PA) column (30 m × 0.32 mm internal diameter) × 1.00 μm df). The GC oven was programmed for 5 min at 200°C, ramped at 10°C/min to 300°C where it was held for an additional 35 min. Carrier gas was helium at a flow rate of 25 ml/min. Compounds were authenticated through comparison of peak retention times with those of purified samples of pear ester and codlemone. Quantification was by integration of peak areas in proportion to that of the internal standard.
Field Studies, 2007–2009.
Studies were conducted in replicated, 0.1–0.2 ha plots (N=5) within mixed-cultivar apple orchards including ‘Delicious,’ ‘Golden Delicious,’ and ‘Granny Smith,’ planted at 370–420 trees ha−1, and with average tree canopies of 3.0–4.5 m. Test orchards were situated near Wapato, WA (46.45° N, 120.42° W) in 2007 and 2009 and near Moxee, WA (46.56° N, 120.39° W) in 2008. Orchards were sprayed uniformly with a number of insecticides for codling moth during each season, including azinphosmethyl (Miles Chemical, Kansas City, MO) and acetamiprid (United Phosphorous, Inc., King of Prussia, PA). Dispenser treatments were randomized at each site and plots were spaced 20–30 m apart. All dispensers were tested at 1,000 ha−1. Dispensers were placed in the upper third of the canopy of each plot. A single delta-shaped trap (Pherocon VI, Trécé Inc.) with a sticky liner and baited with a commercial codlemone lure (Pherocon CM-L2, Trécé Inc.) was placed in the center of each plot and hung at 2.0 m. In addition, five delta-shaped traps with sticky liners and baited with two virgin female codling moths housed inside of a screened plastic cylinder (4.5 cm diameter) hanging from the inside top of the trap were spaced 10–20 m from the center lure-baited trap and placed at 3.0–3.5 m in the canopy. The lure-baited trap at the center of each plot was placed at the lower height to minimize its competitive interaction with female-baited traps.
Studies were divided each year into two time periods that coincided with the first and second generation of codling moth (Knight 2007b). First generation studies were conducted from 16 May to 9 July, 13 May to 30 June, and 5 May to 1 July in 2007, 2008, and 2009, respectively. Studies were continued during the second generation from 16 July to 5 September, 10 July to 8 September, and 7 July to 10 September during these 3 yr, respectively. The codlemone-baited trap was checked and sticky liners were replaced weekly. Lures were replaced after 8 wk. Unsexed moths were collected from the codling moth mass rearing facility at the Yakima Agricultural Research Service Laboratory in Wapato, WA, on Monday of each week, sexed, and stored in plastic crispers at 5°C. Screened cylinders were loaded with two female moths on Tuesday and held at 5°C. Periodic dissections of female moths provided in bulk by this facility have found that <10% of females are previously mated (Knight et al. 2011b). Female-loaded cylinders were taken to the field and loaded in traps on Wednesday of each week. However, only subsets of these data were included in the subsequent analyses. Previous studies using laboratory-reared females have found that moths survive <48 h in these traps if the daily maximum temperatures are >32°C (A.L.K., unpublished data). Data for male catches in the virgin female baited-traps were only included in the analysis when moths were observed to be active for at least 3 d after trap application. This data subset was restricted to male moth catches for 3–7 wk during each generation.
Data were collected and summarized during the two generations (test periods) of codling moth separately. Count and proportional data were transformed (square root and arcsine [square root], respectively) before analysis of variance (ANOVA). Data for the proportion of virgin female traps catching male moths and the cumulative catch of the five virgin female-baited and one lure-baited trap were analyzed with the statistical package, Statistix 9 (Analytical Software, Tallahassee, FL). Differences among means in significant ANOVAs were separated using Tukey's test, P < 0.05. Linear regression was used to compare the daily emission rate of each attractant as a function of their loading at the beginning of each test period. Covariance analysis was used to compare emission rates between each attractant as a function of their initial loading at the start of each time period.
Residual Analysis of Dispensers.
GC analysis of new PVC dispensers found that their actual content was within 20% of the expected loading (Table 1). PVC dispensers field-aged from May to July released a range of 0.42–1.71 and 0.45–1.86 mg/d of codlemone and pear ester, respectively. Mean daily release rates from July to September, however, were lower ranging from 0.16 to 0.54 and 0.05–0.44 mg of codlemone and pear ester, respectively. The linear regressions of mean daily emission of each attractant as a function of the dispenser's initial loading in May and again in July for the second period were significant, P s < 0.0001 with r2s > 0.86 (Fig. 1). The regression's slopes were significantly higher for pear ester versus codlemone during both generations, P < 0.05. The intercepts of the regression lines for each attractant, however, were not different in either the first or second time period, P=0.42 and 0.73, respectively. Mean emission rates of codlemone from Isomate-C Plus dispensers in 2008 and 2009 were lower in the first time period and higher in the second time period with respect to Cidetrak CM (Table 1).
Dispensers were initially placed in the field on 25 May, 12 May, and 28 May, and samples were collected and analyzed on 6 July and 17 Sept., 7 July and 15 Sept., and 8 July and 11 Sept. in 2007, 2008, and 2009, respectively. Isomate-C Plus dispensers were last analyzed in 2008 on 20 Aug.
Field Studies, 2007.
All dispenser-treated plots during the first generation had a significantly lower and similar proportion of virgin female-baited traps catching male moths versus the untreated control (Table 2). Isomate-C Plus and TRE25 (75/0, mg codlemone/mg pear ester, respectively), the codlemone-only PVC dispenser, were not significantly different from the untreated control in the proportion of virgin female-baited traps catching moths in the second generation. The proportion of virgin female-baited traps catching males was significantly lower with all three combo dispensers compared with the untreated control for the second generation. This proportion, however, was significantly lower with TRE24 (45/110) than Isomate-C Plus.
Column means followed by a different letter were significantly different, P < 0.05, Tukey's test.
The cumulative numbers of males caught in both virgin female and codlemone-baited traps in all dispenser treatments were similar and significantly lower than the untreated control in the first generation. In the second generation, all experimental dispensers had similar and significantly lower catches in the codlemone-baited trap than the untreated control. Cumulative catch by the female-baited traps was not significantly different between the untreated control and Isomate-C Plus, but were significantly lower in all the PVC dispenser treatments than in the untreated control. Cumulative catch in female-baited traps was only significantly lower with TRE23 (75/110) than Isomate-C Plus.
Field Studies, 2008.
All PVC combo dispensers had a significantly lower proportion of virgin female-baited traps catching male moths than the untreated control in both generations (Table 3). However, this proportion was not different between the untreated control and Isomate-C Plus in the first generation and between Cidetrak CM and the untreated control in the second generation. Both combo dispensers, TRE144 (45/75) and TRE145 (75/45) had significantly lower proportions of virgin female-baited traps catching males than Isomate-C Plus in the first generation.
Column means followed by a different letter were significantly different, P < 0.05, Tukey's test.
Mean cumulative catch in the female-baited traps were significantly lower with all combo dispensers than in the untreated control in the first generation, while cumulative catches in the virgin female-baited traps were not significantly different among the untreated control and the codlemone-only dispensers Isomate-C Plus and Cidetrak CM. The cumulative catch in female-baited traps during the first generation was significantly lower with TRE144 (45/75) than Isomate-C Plus. Mean cumulative catch of males by female-baited traps though ≈90% lower in dispenser-treated versus untreated plots were not significantly different in the second generation.
Cumulative moth catches in the codlemone-baited trap were significantly lower with all PVC dispensers, but not with Isomate-C Plus than the untreated control during the first generation. Despite 50–95% lower cumulative moth catch in traps baited with the codlemone lure in plots treated with PVC dispensers versus Isomate-C Plus these differences were not significant during the first generation. All dispenser treatments had similar and significantly lower male catches than the untreated control in the codlemone-baited trap during the second generation.
Field Studies, 2009.
No significant difference was found among treatments in the proportion of virgin female-baited traps catching male moths during either the first or second generation (Table 4). However, the two combo dispensers evaluated this year, TRE144 (45/75) and TRE145 (75/45) had the two lowest mean proportions, ≈36–50% lower than the untreated control during the first generation.
Column means followed by a different letter were significantly different, P < 0.05, Tukey's test.
Cumulative moth catches in virgin female-baited traps were significantly lower during the first generation with Cidetrak CM and the combo dispensers TRE144 (45/75) and TRE145 (75/45) than the untreated control. The codlemone-only dispenser, TRE25 (75/0) was not significantly different from the untreated control. Mean catches of males in the virgin female-baited traps were low in the second generation and no difference was found among treatments. Cumulative male catches in the codlemone-baited trap were significantly lower in Cidetrak CM, TRE25 (75/0) and TRE145 (75/45) than the untreated control, and lower in Cidetrak CM than in TRE144 (45/75) during the first generation. Male moth catches in codlemone-baited traps were significantly lower with Cidetrak CM than the untreated check and the combo dispenser TRE144 (45/75) during the second generation.
Pear ester a primary odorant released from ripe pear, Pyrus communis L., elicits a strong attractant response for both larvae and male and female adult codling moth (Knight and Light 2001, Light et al. 2001). Olfactory receptor neurons have been found on male and female antennae that respond specifically to pear ester and others that respond to both pear ester and the sex pheromone, (E,E)-8, 10-dodecadien-1-ol, codlemone (De Cristofaro et al. 2004, Ansebo et al. 2005). Neural processing of the mixed sexual and environmental host plant odor stimuli received by the olfactory receptor neurons in codling moth appears to be through a high level of interactive or across-fiber codling between the glomerular structures of the antennal lobe (Trona et al. 2011). Electrophysiological recordings of glomerular neurons found a range of responses to blends of pear ester and codlemone including synergism and suppression. In addition, specific neurons only responded to the binary blend. This integration of neural processing might be fundamental to the observed increase in behavioral responses of male moths to a blend of codlemone and pear ester reported in field trials (Knight et al. 2005, Villa et al. 2005, Kutinkova et al. 2005, Fernandez et al. 2010) and interactive responses reported in laboratory studies (Yang et al. 2005, Trona et al. 2010).
A number of significant differences were found between dispensers loaded with pear ester and codlemone versus codlemone alone in the disruption of male captures in both virgin female and codlemone-baited traps. Unlike some of the previous studies these differences were found among dispensers all tested at the same density, 1,000 ha−1 and with similar loadings of attractants. In particular, PVC combo dispensers loaded with both codlemone and pear ester outperformed Isomate-C Plus with respect to disruption of virgin female-baited traps. For example, combo dispensers loaded with either 45 or 75 mg pear ester provided significantly greater disruption during the first generation in 2008 and dispensers with 110 mg pear ester were superior in the second generation in 2007 compared with Isomate-C Plus. Unfortunately, during the first generation in both 2007 and 2009 the low number of replicates and high variability among plots contributed to a lack of statistical significance despite increases of 40–50% in the levels of disruption of virgin female-baited traps with combo versus codlemone-only dispensers.
Combo dispensers did not outperform Isomate-C Plus in the disruption of codlemone-baited traps in 2007 and 2008. The only significant difference among dispensers in the disruption of codlemone-baited traps occurred in 2009 when the 120 mg Cidetrak CM dispenser outperformed the combo dispenser loaded with only 45 mg of codlemone (TRE144). Use of lures with lower loadings, such as 0.01–0.03 mg might be a better measure of disruption as the emission rate would be more similar to a virgin female (Bäckman et al. 1997), and would avoid the associated difficulties of using female moths. In addition, the continued use of ≥5 traps baited with the low-load lure per plot is likely an important prerequisite to minimize spatial variability in the analysis of disruption that occurs because of trap placement.
No significant differences were found between the PVC dispensers loaded only with codlemone and Isomate-C Plus. Previous studies have reported that PVC dispensers loaded only with codlemone [TRE9923 (128/0)] disrupted codlemone-baited traps significantly better than Isomate-C Plus during the first generation only (Knight et al. 2011a). Emission rates of codlemone from these two dispensers were found to be statistically similar during the season though TRE9923s was lower during the second generation (Knight et al. 2011a). Similarly, residual analysis of field-aged Isomate-C Plus dispensers in 2008 and 2009 in this study found that this dispenser had a mean daily emission rate of codlemone lower than Cidetrak CM during the first generation and higher in the second generation. Not surprisingly, the level of disruption of virgin female and codlemone-baited traps measured was not statistically different among the PVC dispensers loaded with either 75 or 120 mg codlemone and Isomate-C Plus. These findings are consistent with a previous study of codling moth which found that merely doubling the emission rate of codlemone from two dispenser types applied at the same density (1,000 ha−1) did not affect the disruption of female-baited traps (Knight 1996). However, the two PVC combo dispensers achieved 30 and 50% greater disruption of virgin female-baited traps than Isomate-C Plus during the first generation in 2007 and 2008, respectively.
Unfortunately, our studies have not yet determined an optimal loading of either codlemone or pear ester in PVC dispensers. PVC dispensers loaded with a 10-fold range of pear ester have been field-tested since 2004 (Light et al. 2006, Knight et al. 2011a). Previous results showing that combo dispensers outperform commercial dispensers such as Isomate-C Plus, Isomate-C TT, or Checkmate CM XL1000 were with combo dispensers either loaded at much higher rates or applied at greater densities than the commercial dispensers (Light and Knight 2005, Stelinski et al. 2007, Knight et al. 2011a). Our data reported here are encouraging because combo dispensers loaded with combined rates of pear ester and codlemone that are near equivalent to codlemone loading in commercial dispensers often outperformed these dispensers when applied at the same dispenser density. In particular, the release of pear ester even at subtle emission rates, of 0.1 mg/d−1 during the second generation was seen to numerically increase disruption of virgin female-baited traps.
The relative role of pear ester to codlemone in the combo dispensers is still being resolved. The disruption of male catches in virgin female traps during 2008 was increased >70% in the first generation by substituting pear ester for codlemone, while maintaining the same dispenser density (1,000 ha−1) and total dispenser content (120 mg). This benefit continued in the second generation though reduced to <40% improvement in male disruption. Reductions in the effectiveness of combo dispensers through a season have been reported previously (Steliniski et al. 2007). Analysis of volatiles released by combo dispensers suggested that this pattern was correlated with the differential emission rates of each attractant over time (Knight et al. 2011a). Residual analyses of combo dispensers tested in our study found a significantly higher emission rate of pear ester than codlemone as a function of dispenser content. This differential emission rate was expressed as significant changes in blend ratios emitted by dispensers over the season. For example, maintaining the dispenser loading at 155 mg, but changing the loading ratio of codlemone to pear ester in the 2007 dispensers TRE22 and TRE24 from 2.4:1–1:2.4 altered the emission ratio of the two attractants from 10:1–1:1.7 during the second generation. However, there was no significant difference between dispensers in the disruption of virgin female-baited traps in either generation. Nevertheless, formulating combo dispensers to use the potential benefits provided by pear ester early in the season for codling moth management would most likely be preferred to minimize the reproductive potential of this pest (Solomon 1991).
An alternative approach to use the behavioral effects of pear ester against adult codling moth would be the use of microencapsulated formulation, Cidetrak CM-DA MEC. Applications of this sprayable formulation every 2 wk have enhanced the level of disruption that can be achieved with codlemone-only dispensers (Light and Knight 2005, Knight et al. 2010). Cidetrak CM-DA MEC can also significantly improve the efficacy of some insecticides against codling moth larvae (Schmidt et al. 2008, Light and Beck 2010).
We thank several co-workers at the Agricultural Research Service, USDA in Wapato, WA. Duane Larson and Chey Temple provided technical assistance in the laboratory and field, and Dave Horton provided statistical advice. Helpful reviews of an earlier draft were obtained from Larry Gut, Michigan State University, East Lansing, MI; Greg Krawczyk, Penn State University, Biglerville, PA; and Lukasz Stelinski, University of Florida.