Genotype analysis and studies of pyrethroid resistance of the oilseed rape (Brassica napus) insect pest - pollen beetle (Meligethes aeneus)

摘要

Oilseed Brassicas are vulnerable to attack from many insects and pathogens, calling for an extensive use of pesticides to secure crop yields; this can cause increased resistance in pests. During recent years, one of the main oilseed insect pests—the pollen beetle (Meligethes aeneus), resistant to pyrethroid insecticides—has emerged in southern Sweden. This, because of its frequency and geographic range, provides an excellent source of material for analysis of genetic variation among pollen beetle populations, for study of insecticide resistance and for testing new sources of plant protection. For genotyping pollen beetles, we modified the amplified fragment length polymorphism (AFLP) technique (chosen because it does not depend on prior sequence information when no genome information is available for pollen beetles), and applied it to 133 Swedish populations (susceptible and resistant), collected in different years, and to 14 European populations. AMOVA showed high levels of genetic variation within populations and gene flow among populations, and no evidence of expected regional and resistance-susceptibility to insecticide diversification (clear diversification by time and generations instead) for Swedish populations. European populations showed a clear pattern of regional diversification and a low level of gene flow. To identify possible point mutations associated with pollen beetles resistance to pyrethroids, the primary target sites for pyrethroids—voltage-sensitive sodium channels (VSSC) and metabolic resistance sites—Cytochrome P450, were studied using RT-PCR in resistant and susceptible insects. Two CYP450 partial cDNAs and four cDNA fragments composing VSSC domains I and II were amplified (using primers designed for homologue sequences) and sequenced showing point mutations, which can confer pyrethroid resistance.