Orchard trials in Canada on control ofCydia pomonella (Lep: Tortricidae) by granulosis virus

Cydia pomonella (L.), a major pest of apples in Canadian orchards, is susceptible to a granulosis virus (CpGV). Orchard trials with different formulations of CpGV in Ontario, whereC. pomonella has two generations per year, indicated that as many as seven applications of virus would be needed to maintain the percentage of fruit with deep larval entries below 4%. In Nova Scotia, where there is only one generation ofC. pomonella per year, two applications of CpGV usually were needed although, in some cases, one sufficed. The protection of fruit conferred by CpGV equalled that by the organophosphate insecticides, azinphosmethyl or phosmet, in some tests. The advantages of a selective treatment, such as CpGV, over use of broad-spectrum insecticides are discussed.

---

Résumé Cydia pomonella (L.), un ravageur important du pommier dans les vergers canadiens, est sensible au virus de la granulose. Des essais en vergers utilisant deux préparations de ce virus en Ontario, oùC. pomonella présente deux generations par année, indiquent qu'il faudrait jusqu'à sept pulvérisations pour maintenir à moins de 40% la proportion de fruits portant des infestations larvaires. En Nouvelle Ecosse, oùC. pomonella ne compte qu'une seule génération par an, deux pulvérisations furent nécessaires, quoique dans certains cas une seule a suffi. Dans certains essais, le niveau de protection des fruits égalait celui fourni par les esters phosphoriques, l'azinphos-méthyle ou le phosmet. On discute des avantages des traitements sélectifs tels que le virus de la granulose en comparaison avec l'emploi des insecticides de grande polyvalence.

     

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.     

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.     

Control efficacy of Bacillus thuringiensis and a new granulovirus isolate against Cydia pomonella in orchards

Abstract

Codling moth, Cydia pomonella, is one of the most serious pests of apple and pear worldwide. This study evaluates the efficacies of a granulovirus, a Bacillus thuringiensis (Bt) strain and their combination in the control of C. pomonella in China. A Cydia pomonella granulovirus (CpGV) was isolated from C. pomonella cadavers in an orchard in Gansu, China. Droplet-feeding bioassays showed the median lethal concentration (LC₅₀) of this CpGV isolate (CpGV-C1) against the third instar C. pomonella larvae was 770 OBs µl^?1. The LC₅₀ values of Bt C-33 and kurstaki HD-1 against the third instar larvae were 26.3 µg ml^?1 and 15.7 µg ml^?1, respectively. Field tests indicated the control efficacies of CpGV-C1 and the combination of CpGV-C1 and Bt against C. pomonella larvae in apple orchards were similar to that of beta-cypermethrin. Our data demonstrated that the combination of CpGV and Bt might effectively protect apple fruits from the damage of C. pomonella larvae and had the potential to be developed as a low-cost, highly effective insecticide.     

A model of the effect of codling moth granulosis virus on Cydia pomonella

A model was developed based on an initial ingestion phase, followed by a period of virus multiplication and host death, to describe the effects of codling moth granulosis virus (CpGV) concentration and time from initial infection on the percentage of codling moth larvae (Cydia pornonella) dying from virus infection in the laboratory. With modification, this model also described the effect of CpGV concentration on the reduction of damage to fruit by C. pomonella larvae in the field. In both cases, the effect of CpGV increased with the l/10th power of virus concentration. LT₅₀ (time to death of 50% of larvae) was inversely related to virus concentration in the laboratory. The model predicts the threshold concentration for death of larvae, as well as LD₅₀(C₅₀ and LD₉₀(C ₉₀,) for all ages of larvae. The model also predicts that if the majority of larvae entered through the sides of apple fruits, rather than through the calyx or near the stalk (as they probably did in the field), then the efficacy of CpGV in reducing fruit damage would be lowered. This effect of site of entry is especially marked at low virus concentrations.     

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.     

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.     

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.     

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.     

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.     

Codling moth granulovirus: a comprehensive review

Codling moth (CM), Cydia pomonella (L), is regarded as the most serious insect pest of apple worldwide. A variety of problems associated with the traditional use of non-selective insecticides for its control include: untoward environmental effects, insecticide resistance, negative impacts on natural enemies, and safety for pesticide applicators and the food supply. Concerns about these consequences have increased the interest in and development of alternative means for CM control that have little or no impact on humans, beneficial organisms and sensitive ecosystems. An effective and selective alternative to chemical insecticides for CM control is the CM granulovirus (CpGV). The virus was first isolated in Mexico and subsequently studied and evaluated in Europe and North America. A variety of research including pathology, pathogenesis and histopathology of the virus, determination of virulence, development of production methods, field use, factors that influence efficacy, commercial development, formulation, and CM resistance to the virus has been conducted. Commercial products of CpGV are now produced in Europe and North America and used by orchardists worldwide. In this paper we present a comprehensive review of the CpGV literature and the role of the virus in integrated pest management.     

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.     

Echinococcus multilocularis in north Italy

Alveolar echinococcosis is a zoonotic infection caused by the metacestode of the tapeworm Echinococcus multilocularis. Fox populations living in the Alpine regions of Italy had been considered free from this parasite until 2002, when two infected foxes were detected in Bolzano province (Trentino Alto Adige region) near Austrian border. A modified nested PCR analysis was used to detect E. multilocularis DNA in faecal samples belonging to red fox populations from five Italian regions. A total of 522 faecal samples were analysed from foxes shot in Valle d'Aosta (N = 65), Liguria (N = 44), Lombardy (N = 105), Veneto (N = 67), and Trentino Alto Adige (N = 241) regions. Among these, 24 samples, all from the Trentino Alto Adige region, were found positive. Moreoever, 1406 faecal samples of red foxes were analyzed by CA-ELISAs commercial test kit. This paper provides an update of the epidemiological knowledge of this parasite in north Italy.     

Cydia pomonella granulovirus Genotypes Overcome Virus Resistance in the Codling Moth and Improve Virus Efficiency by Selection against Resistant Hosts

Cydia pomonella granulovirus (CpGV) has been used for 15 years as a bioinsecticide in codling moth (Cydia pomonella) control. In 2004, some insect populations with low susceptibility to the virus were detected for the first time in southeast France. RGV, a laboratory colony of codling moths resistant to the CpGV-M isolate used in the field, was established with collection of resistant insects in the field followed by an introgression of the resistant trait into a susceptible colony (Sv). The resistance level (based on the 50% lethal concentrations [LC₅₀s]) of the RGV colony to the CpGV-M isolate, the active ingredient in all commercial virus formulations in Europe, appeared to be over 60,000-fold compared to the Sv colony. The efficiency of CpGV isolates from various other regions was tested on RGV. Among them, two isolates (I12 and NPP-R1) presented an increased pathogenicity on RGV. I12 had already been identified as effective against a resistant C. pomonella colony in Germany and was observed to partially overcome the resistance in the RGV colony. The recently identified isolate NPP-R1 showed an even higher pathogenicity on RGV than other isolates, with an LC₅₀ of 166 occlusion bodies (OBs)/μl, compared to 1.36 × 10⁶ OBs/μl for CpGV-M. Genetic characterization showed that NPP-R1 is a mixture of at least two genotypes, one of which is similar to CpGV-M. The 2016-r4 isolate obtained from four successive passages of NPP-R1 in RGV larvae had a sharply reduced proportion of the CpGV-M-like genotype and an increased pathogenicity against insects from the RGV colony.     

Horizontal Escape of the Novel Tc1-Like Lepidopteran Transposon TCp3.2 into Cydia pomonella Granulovirus

We characterized an insertion mutant of the baculovirus Cydia pomonella granulovirus (CpGV), which contained a transposable element of 3.2 kb. This transposon, termed TCp3.2, has unusually long inverted terminal repeats (ITRs) of 756 bp and encodes a defective gene for a putative transposase. Amino acid sequence comparison of the defective transposase gene revealed a distant relationship to a putative transposon in Caenorhabditis elegans which also shares some similarity of the ITRs. Maximum parsimony analysis of the predicted amino acid sequences of Tc1- and mariner-like transposases available from the GenBank data base grouped TCp3.2 within the superfamily of Tc1-like transposons. DNA hybridization indicated that TCp3.2 originated from the genome of Cydia pomonella, which is the natural host of CpGV, and is present in less than 10 copies in the C. pomonella genome. The transposon TCp3.2 most likely was inserted into the viral genome during infection of host larvae. TCp3.2 and the recently characterized Tc1-like transposon TC14.7 (Jehle et al. 1995), which was also found in a CpGV mutant, represent a new family of transposons found in baculovirus genomes. The occasional horizontal escape of different types of host transposons into baculovirus genomes evokes the question about the possible role of baculoviruses as an interspecies vector in the horizontal transmission of insect transposons. Received: 27 February 1997 / Accepted: 16 May 1997     

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).     

Transmission of Pepino mosaic virus by the Fungal Vector Olpidium virulentus

Transmission of Pepino mosaic virus (PepMV) by the fungal vector Olpidium virulentus was studied in two experiments. Two characterized cultures of the fungus were used as stock cultures for the assay: culture A was from lettuce roots collected in Castellón (Spain), and culture B was from tomato roots collected in Murcia (Spain). These fungal cultures were maintained in their original host and irrigated with sterile water. The drainage water collected from irrigating these stock cultures was used for watering PepMV‐infected and non‐infected tomato plants to constitute the acquisition–source plants of the assay, which were divided into six different plots: plants containing fungal culture A (non‐infected and PepMV‐infected); plants containing fungal culture B (non‐infected and PepMV‐infected); PepMV‐infected plants without the fungus; and plants non‐infected either with PepMV and the fungus. Thirty‐six healthy plants grouped into six plots, which constituted the virus acquisition–transmission plants of the assay, were irrigated with different drainage waters obtained by watering the different plots of the acquisition–source plants. PepMV was only transmitted to plants irrigated with the drainage water collected from PepMV‐infected plants whose roots contained the fungal culture B from tomato with a transmission rate of 8%. No infection was detected in plants irrigated with the drainage water collected from plots with only a fungus or virus infection. Both the virus and fungus were detected in water samples collected from the drainage water of the acquisition–source plants of the assay. These transmission assays demonstrated the possibility of PepMV transmission by O. virulentus collected from tomato crops.