Codling Moth Granulovirus Research Articles

Biocontrol Potential of the New Codling Moth Granulovirus (CpGV) Strains.

The use of CpGV strains as the basis for bioinsecticides is an effective and safe way to control Cydia pomonella. The research is aimed at the identification and study of new CpGV strains. Objects of identification and bioinformatic analysis: 18 CpGV strains. Sequencing was carried out on a NextSeq550. Genome assembly and annotation were carried out using Spades, Samtools 1.9, MinYS, Pilon, Gfinisher, Quast, and Prokka. Comparative genomic analysis was carried out in relation to the reference genome present in the «Madex Tween» strain-producer (biological standard) according to the average nucleotide identity (ANI) criterion. The presence/absence of IAP, cathepsin, MMP, and chitinase in the genetic sequences of the strains was determined using simply phylogeny. Entomopathogenic activity was assessed against C. pomonella according to the criterion of biological efficacy. Thus, molecular genetic identification revealed that 18 CpGV strains belong to a genus of Betabaculovirus. For all the strains under study ANI values of 99% or more were obtained, and the presence of the cathepsin, chitinase, IAP, and MMP genes was noted. The strains BZR GV 1, BZR GV 3, BZR GV 7, BZR GV 10, and BZR GV L-8 showed the maximum biological efficacy: 100% on the 15th day of observation. Strains BZR GV 4, BZR GV 8, and BZR GV 13 showed efficacy at the level of the «Madex Tween» preparation: 89.5% on the 15th day of observation. The strains with the highest mortality rate of the host insect were identified: BZR GV 9, BZR GV 10, BZR GV L-6, and BZR GV L-8.

Life table simulations of a univoltine codling moth, Cydia pomonella, population 2. Impact of immigration on the effectiveness of codling moth granulovirus sprays

ABSTRACT The potential for immigrant adult codling moths, Cydia pomonella (L.), to reduce the efficacy of the granulovirus (CpGV) against a univoltine population was investigated using published life tables for codling moth on apple trees in Nelson, New Zealand. Simulated life tables over eight generations incorporated mortality >90% from CpGV sprays and combined this with immigration rates ranging from 0 to 1 adult per tree. Without immigration, there were 5 damaged fruits per million by the eighth generation from an initial population of 1.22 adults per tree but only 9 per 100 million had fresh damage or larvae that could trigger phytosanitary rejection. Higher-yielding modern orchards reduced this fresh damage to 5 per 100 million harvested fruit. New Zealand exports apples to markets with zero tolerance of codling moth (Codling Moth Sensitive Markets [CMSM] programme), and immigration of 1 female per 100 trees was projected to pose a risk of failure to meet quarantine requirements with one freshly damaged fruit per 112,000, or per 202,000 in a modern high-density orchard. This confirms the wisdom of using an integrated suite of control methods in CMSM, of which CpGV is only one. Similarly, immigration of one female per 1000 trees or 3 ha had much lower risk but could also threaten exporting if reliance was placed solely on CpGV, with one fresh damage or larva per 2 million harvested fruit. A commitment to the removal of neglected host trees of codling moth in the environs of orchards is recommended.

Compatibility of the egg parasitoid Trichogramma minutum (Hymenoptera: Trichogrammatidae) and the granulovirus (CpGV) used in combination against the codling moth Cydia pomonella (Lepidoptera: Tortricidae)

The codling moth granulovirus (CpGV) and the egg parasitoids of the genus Trichogramma spp. are known as biocontrol agents of codling moth but interactions between them are unknown. The aim of this study was to assess the compatibility and the complementarity of Trichogramma minutum and CpGV. Bioassays were conducted in the laboratory the first year, as well as in an orchard over two years with the two biocontrol agents used alone or in combination. Parasitism of codling moth decreased in glass tube bioassays when CpGV was applied on eggs prior to parasitism, suggesting that female parasitoids discriminated between eggs covered or not with the virus. In contrast, egg discrimination was not observed in the orchard, since the presence of CpGV on codling moth eggs did not affect T. minutum parasitism. Both biocontrol agents together caused 77 and 79% mortality to codling moth in the orchard in the first and the second year of the study, respectively. Our results in the orchard showed that the combined use of CpGV and T. minutum to control the codling moth is compatible. Therefore, the utilization of both biocontrol agents is a promising alternative to control this pest in apple orchards.

Importance of the Host Phenotype on the Preservation of the Genetic Diversity in Codling Moth Granulovirus.

To test the importance of the host genotype in maintaining virus genetic diversity, five experimental populations were constructed by mixing two Cydia pomonella granulovirus isolates, the Mexican isolate CpGV-M and the CpGV-R5, in ratios of 99% M + 1% R, 95% M + 5% R, 90% M + 10% R, 50% M + 50% R, and 10% M + 90% R. CpGV-M and CpGV-R5 differ in their ability to replicate in codling moth larvae carrying the type I resistance. This ability is associated with a genetic marker located in the virus pe38 gene. Six successive cycles of replication were carried out with each virus population on a fully-permissive codling moth colony (CpNPP), as well as on a host colony (RGV) that carries the type I resistance, and thus blocks CpGV-M replication. The infectivity of offspring viruses was tested on both hosts. Replication on the CpNPP leads to virus lineages preserving the pe38 markers characteristic of both isolates, while replication on the RGV colony drastically reduces the frequency of the CpGV-M pe38 marker. Virus progeny obtained after replication on CpNPP show consistently higher pathogenicity than that of progeny viruses obtained by replication on RGV, independently of the host used for testing.

Combining Mutualistic Yeast and Pathogenic Virus — A Novel Method for Codling Moth Control

The combination of a pathogenic virus and mutualistic yeasts isolated from larvae of codling moth Cydia pomonella is proposed as a novel insect control technique. Apples were treated with codling moth granulovirus (CpGV) and either one of three yeasts, Metschnikowia pulcherrima, Cryptococcus tephrensis, or Aureobasidium pullulans. The combination of yeasts with CpGV significantly increased mortality of neonate codling moth larvae, compared with CpGV alone. The three yeasts were equally efficient in enhancing the activity of CpGV. The addition of brown cane sugar to yeast further increased larval mortality and the protection of fruit against larvae. In comparison, without yeast, the addition of sugar to CpGV did not produce a significant effect. A field trial confirmed that fruit injury and larval survival were significantly reduced when apple trees were sprayed with CpGV, M. pulcherrima, and sugar. We have shown earlier that mutualistic yeasts are an essential part of codling moth larval diet. The finding that yeast also enhances larval ingestion of an insect-pathogenic virus is an opportunity for the development of a novel plant protection technique. We expect the combination of yeasts and insect pathogens to essentially contribute to future insect management.

Microencapsulated Pear Ester Enhances Insecticide Efficacy in Walnuts for Codling Moth (Lepidoptera: Tortricidae) and Navel Orangeworm (Lepidoptera: Pyralidae)

The efficacy of combining insecticides with a microencapsulated formulation of ethyl (2E,4Z) -2,4-decadienoate (pear ester, PE-MEC) was evaluated in walnuts, Juglans regia L., for codling moth, Cydia pomonella (L.) (Lepidoptera: Tortricidae), and navel orangeworm, Amyelois transitella Walker (Lepidoptera, Pyralidae). Two types of studies were conducted to compare the use of insecticides with and without PE-MEC. In the first study, PE-MEC in combination with reduced rates of insecticides, including chlorpyrifos, phosmet, methoxyfenozide, and codling moth granulovirus were evaluated in single tree replicates. PE-MEC was tested at one to three rates (0.6, 1.8, and 4.4 g active ingredient ha(-1)) with each insecticide. In the second study, seasonal programs including sprays of esfenvalerate, chlorpyrifos, and ethyl parathion at full rates were evaluated in replicated two ha blocks. Significant reductions in nut injury occurred in the single-tree trial with treatments of PE-MEC plus insecticide compared with the insecticides used alone against both pest species; except with methoxyfenozide for navel orangeworm. Similarly, nut injury in the large plots was significantly reduced with the addition of PE-MEC, except for navel orangeworm in one of the two studies. These results suggest that adding pear ester as a microencapsulated spray can improve the efficacy of a range of insecticides for two key pests and foster the development of integrated pest management tactics with reduced insecticide use in walnut.

Development of a Device to Collect Mass-Reared False Codling Moth,Thaumatotibia leucotreta(Meyrick) (Lepidoptera: Tortricidae), in a Commercial Insectary

The false codling moth, Thaumatotibia leucotreta (Meyrick), has been reared successfully for many years in several small insectaries in South Africa (Schwartz 1971). Rearing was for various purposes, including research, production of the egg parasitoid, Trichogrammatoidea cryptophlebia Nagaraja (Schwartz 1977), and for commercial production of the false codling moth granulovirus, CrleGV (Moore 2002; Moore et al. 2004). The techniques commonly used were adequate for the scale on which rearing was conducted. In 2002 the South African citrus industry initiated research on the sterile insect technique for false codling moth control. As this project progressed it became apparent that the equipment previously used was inadequate for mass rearing false codling moth for commercial sterile insect technique purposes. A programme was launched to design new equipment that would allow for the production of 15 million insects per week. This equipment was installed in a new insectary, Xsit (Pty) Ltd, recently built in Citrusdal, South Africa, for the mass rearing of false codling moth. Amongst others, a moth collection system, using cyclonic separators, was installed. The technical principles in the use of cyclonic separators for particulate matter collection have often been described as in Griffin (1994). However, the development of this type of equipment specifically for the collection of mass-reared moths is poorly documented and often only relates to models suitable for insect collection on a relatively small or laboratory scale, for example Wolf & Stiemann (1971). The use of cyclonic separators in new rearing facilities is therefore probably based on working models in existing mass-rearing facilities.

Effect of Temperature on Long-Term Storage of Codling Moth Granulovirus Formulations

Codling moth, Cydia pomonella (L.), is the major pest of apple (Malus spp.) in the western United States and many other regions of the world. The codling moth granulovirus (CpGV) provides a selective and safe means of its control. We assessed the long-term stability and storage potential of two commercial formulations of CpGV, Cyd-X, and Virosoft. All assays were performed with individual C. pomonella neonate larvae in 2-ml vials on 1 ml of artificial larval diet that was surface inoculated with 10 μl of the test virus suspension. Baseline quantitative assays for the two formulations revealed that the LC50 and LC95 values (occlusion bodies per vial) did not differ significantly between the formulations. For year-long studies on Cyd-X stability, the product was stored at −20, 2, 25, and 35°C, and quantitative bioassays were conducted after 0, 3, 6, and 12 mo of storage. Cyd-X retained good larvicidal activity from −20 to 25°C, and it was the least negatively affected at the lowest temperature. Storage of Cyd-X at 35°C was detrimental to its larvicidal activity within 3 mo of storage. For longer term storage studies, Cyd-X and Virosoft formulations were stored at 2, 25, and 35°C, and assayed for larvicidal activity over a 3-yr period. For recently produced product, a 10-μl sample of a 10−5 dilution of both formulations resulted in 95–100% mortality in neonate larvae. Larvicidal activity for the Cyd-X formulation remained essentially unaffected for 156 wk when stored at 2 and 25°C, but it began to decline significantly after 20 wk of storage at 35°C. The Virosoft formulation stored at 2°C also remained active throughout the 3-yr study, but it began to decline in larvicidal activity after 144 wk at 25°C and 40 wk at 35°C. The information reported in this study should be useful to growers and commercial suppliers for avoiding decreases in CpGV potency due to improper storage conditions.

Effect of Temperature on Long-Term Storage of Codling Moth Granulovirus Formulations

Codling moth, Cydia pomonella (L.), is the major pest of apple (Malus spp.) in the western United States and many other regions of the world. The codling moth granulovirus (CpGV) provides a selective and safe means of its control. We assessed the long-term stability and storage potential of two commercial formulations of CpGV, Cyd-X, and Virosoft. All assays were performed with individual C. pomonella neonate larvae in 2-ml vials on 1 ml of artificial larval diet that was surface inoculated with 10 microl of the test virus suspension. Baseline quantitative assays for the two formulations revealed that the LC50 and LC95 values (occlusion bodies per vial) did not differ significantly between the formulations. For year-long studies on Cyd-X stability, the product was stored at -20, 2, 25, and 35 degrees C, and quantitative bioassays were conducted after 0, 3, 6, and 12 mo of storage. Cyd-X retained good larvicidal activity from -20 to 25 degrees C, and it was the least negatively affected at the lowest temperature. Storage of Cyd-X at 35 degrees C was detrimental to its larvicidal activity within 3 mo of storage. For longer term storage studies, Cyd-X and Virosoft formulations were stored at 2, 25, and 35 degrees C, and assayed for larvicidal activity over a 3-yr period. For recently produced product, a 10-microl sample of a 10(-5) dilution of both formulations resulted in 95-100% mortality in neonate larvae. Larvicidal activity for the Cyd-X formulation remained essentially unaffected for 156 wk when stored at 2 and 25 degrees C, but it began to decline significantly after 20 wk of storage at 35 degrees C. The Virosoft formulation stored at 2 degrees C also remained active throughout the 3-yr study, but it began to decline in larvicidal activity after 144 wk at 25 degrees C and 40 wk at 35 degrees C. The information reported in this study should be useful to growers and commercial suppliers for avoiding decreases in CpGV potency due to improper storage conditions.

Evaluation of the Pear Ester Kairomone as a Formulation Additive for the Granulovirus of Codling Moth (Lepidoptera: Tortricidae) in Pome Fruit

(E,Z) -2,4-decadienoate (pear ester) is a larval kairomone for the codling moth, Cydia pomonella (L.) (Lepidoptera: Tortricidae). Orchard studies were conducted in 2005 and 2006 in apple, Malus domestica Borkhausen, and pear, Pyrus communis L., to evaluate a 5% active ingredient (AI), microencapsulated formulation of pear ester (PE-MEC) as an insecticidal additive for the codling moth granulovirus (CpGV). Although CpGV applied at 5-15-d intervals at commercial rates (2.2 X 10(12)-10(13) granules per ha) killed the majority (82-94%) of larvae found inside infested fruit, it did not eliminate significant damage, i.e., 30-92% fruit injury at harvest versus 51-82% in controls. PE-MEC treatments had significant but inconsistent results in our tests. In apple (mixed cultivars), PE-MEC (3.7-4.7 g [AI] /ha) plus CpGV reduced the percentage of fruit injured during the second but not the first larval generation, compared with CpGV alone, but there no was no additional population reduction (live larvae collected from infested fruit and tree bands). In 'Bartlett' pear, PE-MEC (3.7 g [AI] /ha) plus CpGV significantly increased larval mortality and reduced deep fruit entries at harvest over CpGV alone in 2006, but similar improvements were not observed in 2005 when a lower rate (1.5 g [AI] /ha) was tested. Surprisingly, compared with untreated controls, the PE-MEC formulation alone also reduced fruit injury (mid-season in Bartlett) and larval survivorship inside infested fruit at harvest (2006 apple tests and both years in Bartlett). Although pear ester seems amenable as a kairomonal adjuvant for use with insecticides, our inconsistent data with CpGV in apple and pear suggest practical improvements in formulation and application strategies (e.g., to optimize and maintain attractive release rates) are needed.

Evaluation of the Pear Ester Kairomone as a Formulation Additive for the Granulovirus of Codling Moth (Lepidoptera: Tortricidae) in Pome Fruit

Evaluation of the Pear Ester Kairomone as a Formulation Additive for the Granulovirus of Codling Moth (Lepidoptera: Tortricidae) in Pome Fruit

Evaluation of the codling moth granulovirus and spinosad for codling moth control and impact on non-target species in pear orchards

Abstract We compared the efficacy of a commercial preparation of the codling moth, Cydia pomonella L., granulovirus, CpGV (Cyd-X®) and spinosad (Entrust®) at operational rates for codling moth control in 2004 and 2005. Concurrently we monitored the impact of treatments on populations of non-target arthropods. Spinosad was effective at protecting fruit, with ⩽1.6% codling moth injury in experimental plots, compared with up to 37% injury in the untreated plots at harvest. Mid-season outbreaks of the pear psylla, Cacopsylla pyricola Foerster, were also reduced in spinosad plots. Spinosad was safe for several predators, notably the psylla predator Deraeocoris brevis Uhler, but reduced the abundance of hymenopteran parasitoids by 24% and 40% and non-target Diptera by 49% and 35%, respectively in 2004 and 2005. We found no evidence that spinosad disrupted natural control leading to increased densities of secondary pests including aphids and phytophagous mites. CpGV was less effective than spinosad at protecting fruit, with percentage of fruits attacked similar to controls, but killed the majority (67–71%) of neonate coding moth larvae and did not harm non-target species. Additional observations were conducted in commercial orchards (mixed pear and apple) where CpGV and spinosad were used operationally against existing codling moth infestations. In pear, two spray programs applied in replicated 0.4 ha blocks (i.e. CpGV followed by spinosad against the first and second larval generations, respectively and vice versa) reduced fruit injury at harvest and decreased orchard pheromone monitoring trap catches by 74% over two years. In apple, CpGV was less effective at protecting fruit in the first larval generation compared with spinosad, although population suppression was effective early in the season. Spinosad caused no disruptions of beneficial species or secondary pest outbreaks were observed in the commercial orchards. Our results suggest CpGV and spinosad can be effectively used in integrated pest management for codling moth.

Optimizing Use of Codling Moth Granulovirus: Effects of Application Rate and Spraying Frequency on Control of Codling Moth Larvae in Pacific Northwest Apple Orchards

New formulations of the codling moth, Cydia pomonella (L.), granulovirus (CpGV) [family Baculoviridae, genus Granulovirus] are commercially available in North America. In field tests on apple (Malus sp. ‘Delicious’), we compared different application strategies for CpGV (Cyd-X, Certis USA, Clovis, CA) used in full-season programs against high pest populations. In replicated single tree plots, three rates (0.073, 0.219, and 0.438 liter ha−1) and application intervals (7, 10, and 14 d) killed 81–99% of larvae in fruit and reduced the number of mature larvae recovered in tree bands by 54–98%. Although the proportion of deep entries declined by 77–98%, the amount of fruit injury was not reduced compared with controls. There was a statistical trend between increasing dosage and spray frequency intervals and virus effectiveness, but no interaction between these factors. In a commercial orchard, we assessed a standard (0.219 liter ha−1) and two reduced rates of the virus (0.146 and 0.073 liter ha−1) applied in a weekly spray program in replicated 0.2-ha blocks. In the first generation, fruit injury was reduced in virus-treated compared with three untreated blocks although the decrease was only significant at the standard rate. Mortality rates of larvae (in fruit) were ≥90%, dose dependent, and comparable with rates observed from individual trees sprayed with equivalent treatments in the previous study. Rates of larval mortality declined at all dosages (81–85%) in the first part of the second generation. Most damage and proportionally less mortality occurred in the upper canopy. High pest pressures and untreated blocks contributed to significant damage and the study was terminated early. These data suggest virus programs can be tailored according to the localized pest pressure, but it may not prevent economic damage in high-pressure situations.

Optimizing Use of Codling Moth Granulovirus: Effects of Application Rate and Spraying Frequency on Control of Codling Moth Larvae in Pacific Northwest Apple Orchards

New formulations of the codling moth, Cydia pomonella (L.), granulovirus (CpGV) [family Baculoviridae, genus Granulovirus] are commercially available in North America. In field tests on apple (Malus sp. 'Delicious'), we compared different application strategies for CpGV (Cyd-X, Certis USA, Clovis, CA) used in full-season programs against high pest populations. In replicated single tree plots, three rates (0.073, 0.219, and 0.438 liter ha(-1)) and application intervals (7, 10, and 14 d) killed 81-99% of larvae in fruit and reduced the number of mature larvae recovered in tree bands by 54-98%. Although the proportion of deep entries declined by 77-98%, the amount of fruit injury was not reduced compared with controls. There was a statistical trend between increasing dosage and spray frequency intervals and virus effectiveness, but no interaction between these factors. In a commercial orchard, we assessed a standard (0.219 liter ha(-1)) and two reduced rates of the virus (0.146 and 0.073 liter ha(-1)) applied in a weekly spray program in replicated 0.2-ha blocks. In the first generation, fruit injury was reduced in virus-treated compared with three untreated blocks although the decrease was only significant at the standard rate. Mortality rates of larvae (in fruit) were > or =90%, dose dependent, and comparable with rates observed from individual trees sprayed with equivalent treatments in the previous study. Rates of larval mortality declined at all dosages (81-85%) in the first part of the second generation. Most damage and proportionally less mortality occurred in the upper canopy. High pest pressures and untreated blocks contributed to significant damage and the study was terminated early. These data suggest virus programs can be tailored according to the localized pest pressure, but it may not prevent economic damage in high-pressure situations.

New method for testing solar sensitivity of commercial formulations of the granulovirus of codling moth ( Cydia pomonella, Tortricidae: Lepidoptera)

A method for screening codling moth granulovirus (CpGV) formulation sensitivity to sunlight using specially prepared half apples and a solar simulator is described. The half apple preparation allows an even coverage of virus over the surface of the fruit that would not be possible using whole apples. Leaves and artificial medium were not usable for extended periods of exposure in the solar simulator due to excess drying. Fruit was sprayed with 10 −3 and 10 −5 dilutions of three commercial formulations of CpGV (Carpovirusine, Cyd-X, and Virosoft) and infested with codling moth neonates. Half of the sprayed fruit was exposed to 650 W/m 2 for 4 h in an Atlas Suntest CPS solar simulator resulting in an accumulated radiant energy of 9.36 × 10 6 J/m 2 before they were infested with neonate codling moth larvae. Spraying non-irradiated fruit with the 10 −3 dilution of Cyd-X and Virosoft resulted in nearly 100% mortality of neonate larvae. Irradiation reduced viral activity by 71–98% at the 10 −3 dilution and by up to 32% at the 10 −5 dilution relative to non-irradiated fruit. The procedures utilized enabled good preservation of the fruit throughout the incubation period and minimized invasion of the fruit by plant pathogens and saprophytic organisms. This laboratory method for screening candidate formulations and potential UV protectants could conserve time and resources by eliminating adjuvants with less potential in laboratory tests and field testing only the most promising candidates. It also enables year-round testing.

Efficacy of Codling Moth Granulovirus: Effect of Adjuvants on Persistence of Activity and Comparison With Other Larvicides in a Pacific Northwest Apple Orchard

Control of codling moth, Cydia pomonella (L), in conventional orchards has relied heavily on broad spectrum insecticides such as azinphos-methyl (Guthion®, Bayer CropScience, Research Triangle Park, NC). Alternative control options are needed for a variety of reasons, including environmental impact and worker and food safety concerns. Microbial control agents such as the codling moth granulovirus (CpGV) offer alternatives to conventional insecticides for the control of codling moth. Six weekly applications of the label rate (1 L/ha) of the Carpovirusine® formulation of CpGV in an experimental orchard naturally infested with codling moth provided control of first generation codling moth that was comparable to that of larvicidal oil and azinphos-methyl. Although the number of codling moth entries in fruit that were treated with virus alone was similar to that of control trees, the number of deep entries and the number of living codling moth larvae were significantly reduced on CpGV treated fruit. Despite blemishing, virus-treated fruit with minute entries were suitable for consumption or for processing. Studies on the residual activity of Carpovirusine revealed a steady decline in virus activity 1 to 3 d following application. The use of two adjuvants, Nu-Film-17® and Raynox®, did not protect virus from solar inactivation. Among the biological control options available for codling moth, CpGV provides effective and selective control of neonate larvae. Its use in lieu of broad spectrum insecticides will contribute significantly to the conservation of other natural enemies in the orchard agroecosystem.

Field evaluation of commercial formulations of the codling moth granulovirus: persistence of activity and success of seasonal applications against natural infestations of codling moth in Pacific Northwest apple orchards

Inundative applications of the codling moth (CM), Cydia pomonella L., granulovirus (CpGV), which target neonate larvae before or during initial entry into fruit, offer potential for selective control of this key pest. In field tests on apple we compared the persistence and efficacy of single applications of three CpGV products approved for organic orchards in North America. In addition, the success of repeated (2–14) applications of one product (Cyd-X) as a principal control measure for CM in apple orchards was monitored following operational use by cooperating growers at four separate locations. In the first study, an early season application of all products at label rates remained highly effective for the first 24 h (averaging 94% larval mortality relative to controls) and moderately effective after 72 h (averaging 71% mortality) during dry sunny conditions. Significant activity remained up to 14 days, suggesting prolonged survival of the virus in UV-protected locations, such as the calyx of fruit. A second application later in the season was slightly less effective. Data obtained from commercial sites provide circumstantial evidence for the effectiveness of well-timed CpGV applications against CM outbreaks. In all cases where first generation larvae were targeted beginning at egg hatch (≈250 degree days) and treated areas monitored (0.3–1.6 ha plots), fruit damage during the second larval generation was reduced or eliminated. Based on the number of live larvae recovered throughout the season, mortality rates remained high (80.3–100% across sites). The cumulative number of moths caught in pheromone-baited traps was reduced (66–94%) in the second flight. Data from tree bands placed to catch diapause-destined larvae indicated overwintering generations remained low in treated sites (⩽0.18 larvae/band).