A high-throughput multilocus-amplicon sequencing panel to monitor insecticide resistance in fall armyworm (FAW) Spodoptera frugiperda (Lepidoptera: Noctuidae)

BACKGROUNDFall armyworm (FAW), Spodoptera frugiperda, is a highly polyphagous crop pest that has spread over the world rapidly and invaded Australia in 2020. Globally, FAW has been reported to be resistant to several insecticides permitted in Australia. Timely resistance diagnosis is critical for integrated pest management-based control of FAW in Australia.RESULTSWe developed a multi-amplicon panel by next-generation sequencing (multiamplicon-seq) to identify known insecticide resistance mutations in Australian FAW with high throughput and low cost. The panel included nine known mutations causing insecticide resistance in FAW and four gene mutations causing insecticide resistance in several insect species, not yet reported in FAW. We sequenced 36 plates (96-well) in one MiSeq flow cell with easy sequencing library preparation. We found that Australian FAW carried a very high proportion of the F290V mutation in the acetylcholinesterase (AChE) gene that causes resistance to organophosphate and carbamate insecticides. Furthermore, FAW has a GABA-activated chloride channel mutation, A301Q in the RDL gene. The sequencing-based platform provided evidence of a duplication in the AChE gene. Here several single nucleotide polymorphisms (SNPs) within the 476-bp amplicon of the AChE gene demonstrated 100% heterozygosity across samples and some individuals carried two haplotypes with the F290V mutation.RESULTSWe developed a multi-amplicon panel by next-generation sequencing (multiamplicon-seq) to identify known insecticide resistance mutations in Australian FAW with high throughput and low cost. The panel included nine known mutations causing insecticide resistance in FAW and four gene mutations causing insecticide resistance in several insect species, not yet reported in FAW. We sequenced 36 plates (96-well) in one MiSeq flow cell with easy sequencing library preparation. We found that Australian FAW carried a very high proportion of the F290V mutation in the acetylcholinesterase (AChE) gene that causes resistance to organophosphate and carbamate insecticides. Furthermore, FAW has a GABA-activated chloride channel mutation, A301Q in the RDL gene. The sequencing-based platform provided evidence of a duplication in the AChE gene. Here several single nucleotide polymorphisms (SNPs) within the 476-bp amplicon of the AChE gene demonstrated 100% heterozygosity across samples and some individuals carried two haplotypes with the F290V mutation.RESULTSWe developed a multi-amplicon panel by next-generation sequencing (multiamplicon-seq) to identify known insecticide resistance mutations in Australian FAW with high throughput and low cost. The panel included nine known mutations causing insecticide resistance in FAW and four gene mutations causing insecticide resistance in several insect species, not yet reported in FAW. We sequenced 36 plates (96-well) in one MiSeq flow cell with easy sequencing library preparation. We found that Australian FAW carried a very high proportion of the F290V mutation in the acetylcholinesterase (AChE) gene that causes resistance to organophosphate and carbamate insecticides. Furthermore, FAW has a GABA-activated chloride channel mutation, A301Q in the RDL gene. The sequencing-based platform provided evidence of a duplication in the AChE gene. Here several single nucleotide polymorphisms (SNPs) within the 476-bp amplicon of the AChE gene demonstrated 100% heterozygosity across samples and some individuals carried two haplotypes with the F290V mutation. CONCLUSIONMolecular surveillance by multiamplicon-seq will increase capacity for early detection and future resistance monitoring in highly dispersed Australian FAW. It can provide timely resistance information and has the potential to play an important role in the resistance management of FAW. (c) 2023 Society of Chemical Industry.