Codling moth granulosis virus: effects of its use on some other orchard arthropods

In four field trials from 1978 to 1980, sprays of codling moth granulosis virus (CpGV) plus 1·0% skimmed milk powder did not significantly affect damage to fruit by leaf rollers (tortrix moths). In laboratory tests, survival of larvae of the leaf roller Archips podana fed on leaves sprayed with CpGV plus milk was unaffected and they grew faster than on unsprayed leaves, because of the milk deposits. This might increase damage by A. podana if CpGV plus milk were applied during the feeding period of this species. In one field trial an unusual infestation of fruit by larvae of pith moth Blastodacna atra was not affected by CpGV. Azinphos-methyl significantly reduced damage by B. atru and, in one field trial where sprays were correctly timed, that by leaf rollers. CpGV had no consistently significant effects on numbers of fruit tree red spider mite Panonychus ulmi or its predators, whereas azinphos-methyl induced outbreaks of P. ulmi by killing its predators.     

Key elements in the successful control of diapausing codling moth,Cydia pomonella (Lepidoptera: Tortricidae) in wooden fruit bins with a South African isolate of Heterorhabditis zealandica (Rhabditida: Heterorhabditidae)

The non-insecticidal control strategies currently being implemented in South African orchards for the control of codling moth, Cydia pomonella (L.) may be hampered by wooden fruit bins being infested with diapausing codling moth larvae, acting as a potential source of re-infestation. Key factors contributing to the success or failure of an entomopathogenic nematode application were investigated using the SF 41 isolate of Heterorhabditis zealandica in laboratory bioassays with wooden minibins. Under operational conditions, an application rate of 100 IJs/mL (LD₉₀₌102 IJs/mL) effectively controlled codling moth larvae in these bins, and for further laboratory bioassays, the LD₅₀ value of 18 IJs/mL (?25 IJs/mL) was identified as the discriminating dosage. Maximum mortality was attained when bins were pre-wet for at least 1 min (>90% RH) and maintained at maximum humidity (>95% RH) post-treatment for at least 3 days (LT₉₀=73 h), to ensure nematode survival and subsequent satisfactory infection of diapausing codling moth larvae. Tarping bins achieved the desired high level of humidity required. Furthermore, adjuvants (specifically Reverseal 10?) also improved an application. The study conclusively illustrated that if all the above-mentioned conditions are met, H. zealandica has the potential to successfully disinfest wooden fruit bins of codling moth.     

The Role of Formulation Additives in Increasing the Potency of Cydia pomonella Granulovirus for Codling Moth Larvae,in Laboratory and Field Experiments

Studies were undertaken to improve the biological efficacy of the granulovirus (CpGV) of the codling moth, Cydia pomonella , by evaluating the performance of some formulation additives that might improve virus persistence and/or virus uptake by first instar larvae. Laboratory studies, using a leaf disc bioassay, demonstrated that 15% cane molasses incorporated within a formulation of purified CpGV dramatically reduced the median lethal exposure time (LET 50 ) to CpGV for neonate larvae at a CpGV dosage rate of 10 7 occlusion bodies (OBs) ml -1 . Screening of a range of other compounds showed that sucrose, fructose and sorbitol (at 10% concentrations) and extracts of apple flesh and skin also gave significant reductions in the LET 50 of CpGV formulations containing these ingredients. Pectin, malic acid and &#102 -farnesene did not significantly reduce the LET 50 . In a field trial, molasses included at 15% (v/v) in a CpGV formulation, containing a dosage rate of 10 12 OBs ha -1 , gave as good control of codling moth damage as virus formulations containing the 'sticker' 0.2% skimmed milk at higher dosage rates of 10 13 and 10 14 OBs ha -1 . Studies of CpGV persistence on foliage revealed no significant improvement of virus persistence on apple foliage using 10% or 15% molasses formulations. A second field trial demonstrated that 10% molasses, 10% sorbitol or 0.08% &#102 -farnesene significantly reduced codling moth deep damage to fruit when these ingredients were added to formulations of pure CpGV. Substantial sooty-mould growth ( Cladosporium spp.) was observed on apple foliage treated with formulations containing molasses, indicating that this formulation additive has secondary consequences that would need to be taken into account if molasses was to be used in commercial CpGV formulations. Nonetheless, these studies clearly demonstrate that major biological improvements in CpGV performance can be achieved by the incorporation of formulation additives, including molasses and several other compounds, that probably function as attractants and/or feeding stimulants for codling moth larvae.     

Characterisation of Cydia pomonella granulovirus from codling moths in a laboratory colony and in orchards of British Columbia*

Sequence analyses indicate that a granulovirus isolated from a laboratory colony of codling moth in British Columbia, Canada is identical or very similar to the Mexican isolate of Cydia pomonella granulovirus (CpGV-M). In addition to codling moth, the vims was found to cause mortality of secondary leafroller pests of North American orchards, including the fruit tree leafroller and the obliquebanded leafroller. A survey in apple growing regions of British Columbia indicates that 23% of the wild population of codling moth larvae carry CpGV.     

Comparative activity of baculoviruses against the codling moth Cydia pomonella and three other tortricid pests of tree fruit

The granulovirus of Cydia pomonella (L.) (CpGV) offers potential for selective control of codling moth. Two major limitations of CpGV are its narrow host range and lack of persistence in the orchard agroecosystem. The nucleopolyhedroviruses of the alfalfa looper Autographa californica (Speyer) (AcMNPV) and those of the celery looper Anagrapha falcifera (Kirby) (AfMNPV) have broad host ranges. Comparative assays of CpGV, AcMNPV, and AfMNPV against codling moth neonate larvae revealed a 54-93-fold greater susceptibility of codling moth to the granulovirus than to the two nucleopolyhedroviruses based on the LC(50) values for each virus. The LC(50)s for CpGV, AfMNPV, and AcMNPV were 32.7 capsules/mm(2), 1.77 x 10(3) occlusion bodies (OBs)/mm(2), and 3.05 x 10(3)OBs/mm(2), respectively. The LT(50) determined for AfMNPV using an approximate LC(95) of the virus against neonate larvae was 3.6 days. Histological examination of tissues in moribund codling moth larvae that had been treated with AfMNPV revealed the presence of nonoccluded and unenveloped virus rods in midgut tissue. Neither OBs nor signs of infection were detected in other tissues. The activity of AfMNPV was also evaluated in three other tortricid apple pests (obliquebanded leafroller, Choristoneura rosaceana (Harris); Pandemis leafroller, Pandemis pyrusana Kearfott; and the oriental fruit moth, Grapholitha molesta (Busck)). Codling and Oriental fruit moths were significantly more susceptible to AfMNPV than were the two leafroller species.     

The susceptibility of the pea moth, Cydia nigricana, to infection by the granulosis virus of the codling moth, Cydia pomonella

Laboratory studies demonstrated that neonate larvae of the pea moth, Cydia nigricana, are susceptible to infection with a granulosis virus (CpGV) isolated from the codling moth, Cydia pomonella. Comparative LC₅₀ values for C. nigricana and C. pomonella are 1.90 × 10⁵ and 1.54 × 10⁴ capsules/ml of diet, respectively. The virus extracted from CpGV-infected pea moth larvae is serologically related, and probably identical, to CpGV.     

Development of a direct test of baculovirus resistance in wild codling moth populations

The occurrence of resistance of codling moth (CM, Cydia pomonella L.) to Cydia pomonella granulovirus (CpGV) used as biological control agent revealed the need for fast and reliable resistance monitoring methods. Here, we describe the development of a laboratory resistance test that is directly performed on larvae extracted from infested apples. This test is based on a 14‐day bioassay at a discriminative CpGV concentration of 2 × 10⁵ occlusion bodies/ml diet and can be applied to L1–L4 larvae. Information on virus resistance can be obtained within <4 weeks. In a survey, CM larvae were isolated and tested from 6698 apples from 10 different orchards in Germany, Austria, Switzerland, Italy and the Netherlands. We identified seven CM populations resistant or partly resistant to CpGV isolates. Although some of the orchards were treated with commercial CpGV products, this method allowed us to obtain reliable information about the resistance status of the examined populations.     

Field trials of CpGV virus isolates overcoming resistance to CpGV-M

The Cydia pomonella granulovirus (CpGV) has been used for many years as biological agent for codling moth control in apple orchards. Resistance to the Mexican strain of CpGV was detected in orchards in Germany, France and Italy. A laboratory insect colony was started from insects collected in a French resistant orchard. It was named RGV. Various virus isolates were identified as active against this resistant insect colony. Field tests were carried out in 2007 to test if the two virus isolates CpGV-I12 and NPP-R1 were effective in the field. Although these virus isolates were not able to reduce insect caused fruit damages, they significantly reduced the overwintering insect populations. NPP-R1 was subjected to eight passages on RGV larvae (NPP-R1.8) that improved its biological activity on RGV larvae. 2008 field trials were set up to test this improved virus strain, compared to CpGV-I12 and Madex plus active on RGV. These tests confirmed the ability to control both in susceptible and resistant insect populations.     

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/m² for 4 h in an Atlas Suntest CPS solar simulator resulting in an accumulated radiant energy of 9.36 × 10⁶ J/m² 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.     

Investigating the horizontal transmission of the Cydia pomonella granulovirus (CpGV) in a model system

Repeated applications of Cydia pomonella granulovirus (CpGV) can effectively control the codling moth (CM) in apple orchards. However, it is still unknown whether horizontal transmission of the virus from infected to uninfected larvae contributes to the efficacy of the virus insecticide. Horizontal transmission of CpGV was assayed using detached apples. In experiments using artificially applied virus dots on the apple’s surface or infected CM larvae as virus inoculum, it was found that the likelihood of infection of healthy CM larvae relied mainly on the larval behavior. The amount of virus inoculum, either applied artificially or produced by the infected larvae, impacted the infection rate only to a small degree. In the experiments, CM larvae exhibited a strong preference in entry sites, increasing the chance for horizontal transmission. Depending on the experimental design, horizontal transmission rates of about 40% were observed in laboratory assays.     

The occurrence and distribution of resistance of codling moth to Cydia pomonella granulovirus in Europe

Thirty‐five codling moth (CM, Cydia pomonella L., Lepidoptera, Tortricidae) populations collected in different commercial orchards in six European countries were tested for their susceptibility to Cydia pomonella granulovirus (CpGV‐M). Including previously published data on CpGV‐M resistance, a total of 38 CM colonies showed considerably elevated LC₅₀ values, independent of the country origin. When only few test individuals are available, determination of mortality of neonate larvae at a discriminating concentration range of 10⁴ to 10⁶ OB/ml (>log4) as a direct measure of percentage susceptible individuals in a CM population is more advisable than calculation of LC₅₀ values. The >log4 mortality alone or in combination with the LC₅₀ value can be used for identification of resistance in a population. Results indicated a locally separated but widely spread occurrence of CM populations with low susceptibility to CpGV‐M. The most plausible hypothesis for the emergence of CpGV‐M resistance is its selection by repeated use of CpGV products.     

Host range extension of <Emphasis Type="Italic">Cydia pomonella</Emphasis> granulovirus: adaptation to Oriental Fruit Moth, <Emphasis Type="Italic">Grapholita molesta</Emphasis>

Among various Cydia pomonella granulovirus (CpGV) isolates, the Mexican isolate (CpGV-M) has demonstrated a significant ability to reduce damage induced by the oriental fruit moth, Grapholita molesta (Busck) (=Cydia molesta) in peach crops. To obtain a more efficient virus for G. molesta control, an experimental virus population was constructed by mixing various CpGV isolates. This mixture was then selected for replication in a G. molesta laboratory colony. After 12 successive passages on this alternative host, the insecticidal efficacy of the virus population had improved. The concentration of virus occlusion bodies required to kill 90 % of neonate larvae was 450-fold lower than that of the original isolate mixture, and 120-fold lower than that of the CpGV-M isolate alone. Following adaptation to this alternative host, the efficacy against its natural host, the codling moth, C. pomonella, was conserved. This mixed isolate population can be produced on C. pomonella without loss of efficacy, which is useful from a commercial production perspective. This adapted virus isolate mixture is likely to prove more effective than individual component isolates at controlling G. molesta.     

Incidence and transmission of a granulovirus in a large codling moth [Cydia pomonella L. (Lepidoptera: Tortricidae)] rearing facility

Incidences of potential per os Cydia pomonella granulovirus (CpGV) transmission within a large codling moth colony were identified. CpGV was detected in the water which is used to wash egg sheets. When pre-neonates were extracted from eggs prior to emergence and tested for the presence of CpGV, 40% were found to carry amounts of CpGV detectable by a polymerase chain reaction (PCR) assay, suggesting possible transovarial transmission of the virus. Although symptoms typical of virus infection and larval death were found infrequently within communal rearing trays, the frequency with which CpGV DNA was detected by PCR assays increased from a mean of 31% of 10-day-old larvae to 94% of 25-day-old larvae. CpGV in codling moth cadavers remained virulent after being held at 60 degrees C for 3 days under conditions similar to the treatment of spent diet at the rearing facility before its disposal. PCR tests of surface samples taken from air filters and rearing rooms of the rearing facility were found to contain CpGV. Bioassays of surface samples from the diet trash bin and a filter through which outside air is passed before entering the rearing chambers resulted in significant codling moth neonate mortality. The virulence of CpGV in dust from the spent diet and the original inadvertent positioning of the diet trash bin directly below one of the air intake ducts are suggested as a possible additional source of CpGV contamination within the facility.     

Uptake of Granulovirus from the Surface of Apples and Leaves by First Instar Larvae of the Codling Moth Cydia pomonella L. (Lepidoptera: Olethreutidae)

Cydia pomonella granulovirus (CpGV) has received considerable attention as a potential microbial insecticide for the control of the codling moth (Cydia pomonella L.) , a worldwide pest of apples. Laboratory experiments were established to investigate virus uptake by first instar larvae, using a novel leaf disc bioassay technique. Virus uptake was found to be independent of active feeding and larvae became infected simply by walking or browsing on sprayed leaf disc surfaces in as little time as 3.5 min. Infection increased as a function of time spent on the leaf disc surface and a linear log time/probit mortality relationship could be fitted. The bioassay technique used has potential for the realistic laboratory testing of virus spray formulations. A field experiment showed that virus infection could be contracted by newly hatched codling moth larvae both from the surface of sprayed leaves and sprayed fruit. The potential for exploiting this knowledge for improving spray formulations is discussed.     

Evaluation of lignins and particle films as solar protectants for the granulovirus of the codling moth,Cydia pomonella

The identification of effective solar protectants for field application of the codling moth, Cydia pomonella (L.), granulovirus (CpGV) is of interest to improve its efficacy and commercial viability as a biological pesticide. We evaluated several materials as potential adjuvants to protect CpGV from ultraviolet degradation. In laboratory tests with a solar simulator (9.36×10⁶ J/m²), the addition of kaolin clay (Surround® WP at 3 and 6%, w/v), a paraffin wax-based emulsion (SPLAT? at 5%, v/v), and a bark extract trans-cinnamaldehyde combined with a film agent (both at 1%, v/v) did not significantly reduce larval entries or increase larval mortality in irradiated apples that were treated with a commercial CpGV product (Cyd-X). In semi-field tests in an apple orchard, a spray-dried lignin formulation containing CpGV (6.57×10¹² OBs/ha) and a lignin-based adjuvant used with Cyd-X (both applied at 4.7–5.6 kg lignin/ha) significantly improved residual activity of CpGV compared with Cyd-X alone applied at the same rate. However, the benefits were short lived and could not be detected after 7 days. In orchard tests, we evaluated two additional refined lignin-products (Lignosulfonate and Vanisperse? CB at 5.61 kg/ha) and two particle film materials (kaolin clay, ‘Cocoon?’, and calcium with boron ‘Eclipse?’) as adjuvants for UV protection of Cyd-X (6.57×10¹² OBs/ha) in tests against a dense codling moth infestation. Although all virus treatments were highly effective (causing ≥90% larval mortality), no significant effects of the adjuvant treatments could be detected. In these latter tests, the use of a silicone based wetting agent at 0.025% (v/v) may have been beneficial at increasing mortality among older larvae inside the fruit.     

Ultraviolet protection of the Cydia pomonella granulovirus using zinc oxide and titanium dioxide

Cydia pomonella granulovirus (CpGV) is a specific pathogen of codling moth, the most serious pest of apple worldwide and has recently been isolated in China. However, its use for codling moth control is limited by ultraviolet (UV) solar radiation, which is a major factor affecting the field persistence of this virus. The virion is occluded in the granulin matrix of occlusion bodies. Many substances have been tested as sunscreen agents, but little has been published on the use of reflectors with the occluded bodies (OBs) of CpGV. This work investigates the susceptibility of a native GV, CpGV-ZY, to UVB radiation over different time periods and evaluates the protective effect of two sunscreen agents, zinc oxide (ZnO) and titanium dioxide (TiO₂). Laboratory tests showed 10⁴ OB/ml of CpGV-ZY exposed to UVB light (3.5 W/m²) for 3.75 h caused 50% inactivity. At 15 mg/ml ZnO and 10 mg/ml TiO₂, the mortality was highest after 4-h exposure to UVB light. Semi-field tests indicated both compounds are effective as UV protectants at low concentrations. These are the first results confirming that ZnO and TiO₂ hold promise as UV protectants for this CpGV-ZY isolate. Moreover, it is apparently safe and effective to use within the range of concentrations needed for codling moth control.     

Dosage-mortality and time-mortality studies of a granulosis virus in a laboratory strain of the codling moth,Laspeyresia pomonella

Different doses of a granulosis virus were administered to first- and fifth-instar larvae of the codling moth Laspeyresia pomonella. Virus was very pathogenic for both larval instars. The LD₅₀ values for first- and fifth-instar larvae were 5 and 49 capsules/larva, respectively. However, fifth-instar larvae were much more variable in their response to virus than first-instar larvae. Using probit methods it was calculated that 1 capsule could cause death in about 25% of both larval instars but 1578 capsules were required to cause 70% mortality of fifth-instar larvae as compared to 12 capsules for first-instar larvae. This is the first report of a decided difference in variability of response to virus by two larval instars of the same species. A bimodal response by both larval instars was observed in time-mortality studies. Apparently, about 20% of the larvae were very resistant to virus infections.     

苹果蠹蛾抗药性研究进展

苹果蠹蛾是世界各国高度关注的严重危害苹果生产的外来有害生物。该虫于20世纪50年代在我国首次报道,目前是我国一类进境检疫性有害生物,正严重威胁我国苹果主产区的水果生产安全。苹果蠹蛾以幼虫钻蛀到果实内部为害,防治难度高,对其主要采用化学农药、交配干扰和苹果蠹蛾颗粒体病毒进行防治。由于农药的长期大量使用,苹果蠹蛾已对有机磷、氨基甲酸酯、拟除虫菊酯、昆虫生长调节剂、阿维菌素和苹果蠹蛾颗粒体病毒等不同类型的杀虫剂产生了抗药性。本文总结了国内外有关苹果蠹蛾抗药性现状和抗药性机理方面的研究,并分析了其对几种农药产生抗性的主要原因,同时结合国外苹果蠹蛾防治和抗药性相关研究,以及其在我国发生与防治的现状,提出该虫抗药性治理策略,即及时对我国疫区苹果蠹蛾的抗药性现状进行监测,在此基础上,注意科学地使用化学农药,并结合农业防治和生物防治等措施对该虫进行综合治理。     

Overcoming the resistance of codling moth against conventional Cydia pomonella granulovirus (CpGV-M) by a new isolate CpGV-I12

Recently, codling moth (CM, Cydia pomonella L.) populations with a significantly reduced susceptibility to C. pomonella granulovirus (CpGV) products have been observed in Germany. A novel CpGV isolate, designated CpGV-I12, is able to overcome the CpGV resistance. CpGV-I12 originated from Iran and showed superior efficacy in laboratory bioassays against a resistant CM strain (CpR), which has a 100-fold reduced susceptibility to commercially used isolate CpGV-M. Determination of the median lethal concentration (LC₅₀) indicated that CpGV-I12 is nearly as efficient in resistant CpR as CpGV-M in a susceptible CM strain (CpS). Beyond, CpGV-I12 caused superior mortality in CpS. Infection experiments showed that the resistance breaking effect can be observed in all instars of CpR. CpGV-I12 is a promising alternative control agent of CM in orchards where conventional CpGV products fail. In addition, we demonstrate in bioassays with recombinant expressed Cry1Ab that cross-resistance to CpGV and Bacillus thuringiensis products is not likely.     

Production and field evaluation of codling moth granulosis virus for control of Cydia pomonella in the United Kingdom

Codling moth granulosis virus (Cp GV) was produced in larvae of Cydia pomonella reared on artificial diet. The average yield of virus (9 × 10⁹capsules/larva) was increased by raising the larvae on diet containing methoprene, a juvenile hormone analogue. In field trials in 1978 and 1979, one or two high-volume applications of Cp GV at 7 × 10¹⁰ capsules/litre achieved reductions in numbers of mature larvae and damaged fruit that were little different from those obtained by two applications of azinphos-methyl. A field trial in 1980 showed that at concentrations of Cp GV ≥ 6 × 10⁸ capsules/litre, damaged diminished slowly with increasing virus concentration. Following virus application in 1980, Cp GV infectivity was reduced by half in 3 days, but some activity persisted at least 4–8 wk after spraying. The results indicate that codling moth GV is an effective control agent particularly in reducing numbers of larvae and the more severe forms of fruit damage.