High resolution melting point application to the detection of the relative frequencies of genotypes in mixed infections of CpGV in coding moth

Abstract

International audienceThe High resolution melting (HRM) was adapted to discriminate between two CpGV genotypes that can be present together in treated apple orchards: CpGV-M and CpGV-R5. CpGV-M is the prototype of the Cydia pomonella granulovirus. When a larva carrying the type I resistance ingest CpGV-M alone, the virus replication is blocked. CpGV-R5 is a virus isolate which replication is not affected by the presence of the type I resistance; it replicates similarly in insects carrying or not the type I resistance gene. When both genotypes are present in the same larva, both replicate, and WKH HIÀFDF\ RI FRQWURO LV KLJKHU ,W LV LPSRUWDQW WR GHWHUPLQH ERWK WKHsuccess of replication in larvae taken from treated orchards, and the relative proportions of each virus genotype, to assess the resistance levels of the insect population. The HRM method takes advantage of 24 bps size difference between the pe38 gene alleles present in CpGV-R5 and CpGV-M. The allele present in CpGV-R5 is shorter compared with that in CpGV-M. The method has been applied to the detection of genetically pure and mixed viral populations. It has been tested on mixtures of Occlusion Bodies, and also on OBs recovered from larvae dying from mixed infections. Last, tests were done using hemolymph -that contains Budded Virus- from infected larva. Samples containing 90% of one genotype and 10% of the second are reliably qualified