Novel types of resistance of codling moth to Cydia pomonella granulovirus

The Cydia pomonella granulovirus (CpGV, Baculoviridae) is an important biological control agent to control codling moth (CM; Cydia pomonella, L.) in organic and integrated pome fruit and walnut production. The CpGV is highly host-specific and supremely virulent for early larval stages of CM, additionally safe for the environment and other animals and humans. Since 2005, resistance against the widely used Mexican isolate (CpGV-M) has been reported from different countries in Europe. Until now, over 40 apple orchards with resistance to CpGV-M based products were identified. For several CM field populations in Europe a Z-linked, monogenetic and dominant inheritance was proposed suggesting a highly similar mode of resistance, termed type I resistance. Type I resistance is targeted only against the isolate CpGV-M and specific the 24 bp insertion in its viral gene pe38. Some other CpGV isolates collected from infected larvae of different geographical regions, lacking this 24 bp repetitive insertion in their pe38 gene and caused virus infection in resistant larvae. Some of these isolates, e.g. CpGV-S, were eventually registered to re-establish the efficient control of CM larvae in the field. Recently, two CM field populations, called NRW-WE and SA-GO, with an untypically high resistance level against CpGV-M and other CpGV isolates, were identified and a novel resistance type II was proposed. This thesis focuses on the elucidation of their mode of inheritance and their cellular mechanism. For generating genetically homogenous resistant strains out of the field population NRW-WE, larvae were selected by repeated mass crosses and selection under virus pressure, using the two isolates CpGV-M and CpGV-S, respectively. The resulting strains CpR5M and CpR5S showed a clear cross-resistance to both CpGV-M and CpGV-S. By crossing and backcrossing experiments between CpR5M or CpR5S and susceptible CM strain (CpS) an autosomal dominant and monogenetic inheritance of resistance was elucidated. The autosomal inheritance mode supported the evidence of a second type (type II) of resistance. Initially, an interchromosomal rearrangement involving the Z chromosome was hypothesized to explain the translocation from a Z-chromosomal to an autosomal inheritance. This hypothesis, however, could be clearly ruled out because a highly conserved synteny of all probed Z-linked genes was observed for different resistant CM strains when fluorescence in situ hybridization with marker genes (BAC-FISH) was applied. Considering the cross-resistance in type II resistance, CM larvae were treated with single or mixtures of the isolates CpGV-M and CpGV-S. For these treatments no virus infection was observed but a recombinant of CpGV-M containing the pe38 gene of CpGV-S caused high mortality. The results indicated that beyond the known pe38 related mechanism of type I resistance against CpGV-M, a second mechanism seemed to exist in type II resistance. With CpR5M and CpS budded viruses injections, circumventing initial midgut infection, gave further evidence that resistance against CpGV-S is midgut-related. A fluoresecence-quenching assay using rhodamin-18 labeled occlusion derived viruses could not fully elucidate whether receptor binding or an intercellular midgut factor is involved in type II resistance. The results led to the model of two different but genetically linked resistance mechanisms in the type II resistant CM larvae: resistance against CpGV-M is systemic and targeted against the pe38 gene, whereas resistance against CpGV-S is based on an unidentified midgut factor, inhibiting initiation of infection. A further CM field population, termed SA-GO, was also investigated for the biological and genetic background of CpGV resistance. Crossing experiments between CpS and field collected larvae of SA-GO, followed by resistance testing with two CpGV isolates revealed differences in the susceptibility and the mode of inheritance compared to the one found in type I or type II resistance of CM. Single-pair inbreeding generated the genetically more homogenous resistant strain CpRGO. Reciprocal hybrid crosses and backcrosses between individuals of CpRGO and susceptible CpS observed a dominant and polygenic inheritance of resistance in the majority of crosses. Resistance to CpGV-S appeared to be autosomal and dominant for larval survivorship but recessive when success of pupation of the hybrids was considered. Resistance of CpRGO to CpGV-M however, is proposed to be both autosomal and Z-linked inherited, since only male larvae were able to pupate, similar to the type I resistance. CpRGO was therefore termed type III resistance. When the efficacy of different CpGV isolates classified to all known CpGV genome groups (A - E) was tested with neonates of all resistant strains. CpGV isolates of the genome groups B and C were able to cause significant mortality in larvae of all resistance types. In addition, CpGV of genome group D caused high mortality in type III resistant CM strain, whereas type I resistance was broken by all known CpGV genome groups, except group A. When isolates of commercial CpGV products were tested in the resistant CM strains, it was found that the commercially used CpGV isolates R5 and 0006 did break only type I and type III resistance, whereas isolate V15 was able to cause high mortality in all resistant types. In conclusion, two types of CpGV resistance, type II and type III were identified and showed a high heterogeneity in their mode of inheritance, mode of action and response to CpGV isolates of different genome groups. The major finding of this thesis is that field resistance of CM to CpGV is genetically and functionally variable and needs to be carefully addressed when resistance management strategies are developed for CM control in the field.