title: Ryanodine receptor point mutations confer diamide insecticide resistance in tomato leafminer, Tuta absoluta (Lepidoptera: Gelechiidae).
abstract: Insect ryanodine receptors (RyR) are the molecular target-site for the recently introduced diamide insecticides. Diamides are particularly active on Lepidoptera pests, including tomato leafminer, Tuta absoluta (Lepidoptera: Gelechiidae). High levels of diamide resistance were recently described in some European populations of T. absoluta, however, the mechanisms of resistance remained unknown. In this study the molecular basis of diamide resistance was investigated in a diamide resistant strain from Italy (IT-GELA-SD4), and additional resistant field populations collected in Greece, Spain and Brazil. The genetics of resistance was investigated by reciprocally crossing strain IT-GELA-SD4 with a susceptible strain and revealed an autosomal incompletely recessive mode of inheritance. To investigate the possible role of target-site mutations as known from diamondback moth (Plutella xylostella), we sequenced respective domains of the RyR gene of T. absoluta. Genotyping of individuals of IT-GELA-SD4 and field-collected strains showing different levels of diamide resistance revealed the presence of G4903E and I4746M RyR target-site mutations. These amino acid substitutions correspond to those recently described for diamide resistant diamondback moth, i.e. G4946E and I4790M. We also detected two novel mutations, G4903V and I4746T, in some of the resistant T. absoluta strains. Radioligand binding studies with thoracic membrane preparations of the IT-GELA-SD4 strain provided functional evidence that these mutations alter the affinity of the RyR to diamides. In combination with previous work on P. xylostella our study highlights the importance of position G4903 (G4946 in P. xylostella) of the insect RyR in defining sensitivity to diamides. The discovery of diamide resistance mutations in T. absoluta populations of diverse geographic origin has serious implications for the efficacy of diamides under applied conditions. The implementation of appropriate resistance management strategies is strongly advised to delay the further spread of resistance.
authors.affiliation: Hellenic Agricultural Organisation - 'Demeter', Institute of Olive Tree, Subtropical Crops and Viticulture, Heraklion, Crete, Greece. Electronic address: eroditakis@nagref.gr.
authors.forename: Emmanouil
authors.initials: E
authors.lastname: Roditakis
authors.affiliation: Bayer CropScience, R&D Pest Control, Monheim, Germany; Department of Biology, Martin-Luther-Universität Halle-Wittenberg, Halle, Germany.
authors.forename: Denise
authors.initials: D
authors.lastname: Steinbach
authors.affiliation: Department of Biology, Martin-Luther-Universität Halle-Wittenberg, Halle, Germany.
authors.forename: Gerald
authors.initials: G
authors.lastname: Moritz
authors.affiliation: Hellenic Agricultural Organisation - 'Demeter', Institute of Olive Tree, Subtropical Crops and Viticulture, Heraklion, Crete, Greece.
authors.forename: Emmanouil
authors.initials: E
authors.lastname: Vasakis
authors.affiliation: Hellenic Agricultural Organisation - 'Demeter', Institute of Olive Tree, Subtropical Crops and Viticulture, Heraklion, Crete, Greece.
authors.forename: Marianna
authors.initials: M
authors.lastname: Stavrakaki
authors.affiliation: Hellenic Agricultural Organisation - 'Demeter', Institute of Olive Tree, Subtropical Crops and Viticulture, Heraklion, Crete, Greece.
authors.forename: Aris
authors.initials: A
authors.lastname: Ilias
authors.affiliation: Departamento de Producción Vegetal, Universidad Politécnica de Cartagena, Cartagena, Spain.
authors.forename: Lidia
authors.initials: L
authors.lastname: García-Vidal
authors.affiliation: Departamento de Producción Vegetal, Universidad Politécnica de Cartagena, Cartagena, Spain.
authors.forename: María Del Rosario
authors.initials: MDR
authors.lastname: Martínez-Aguirre
authors.affiliation: Departamento de Producción Vegetal, Universidad Politécnica de Cartagena, Cartagena, Spain.
authors.forename: Pablo
authors.initials: P
authors.lastname: Bielza
authors.affiliation: Institute of Molecular Biology & Biotechnology, Foundation for Research & Technology Hellas, Crete, Greece.
authors.forename: Evangelia
authors.initials: E
authors.lastname: Morou
authors.affiliation: Departamento de Agronomia, Universidade Federal Rural de Pernambuco - UFRPE, Recife, Brazil.
authors.forename: Jefferson E
authors.initials: JE
authors.lastname: Silva
authors.affiliation: Departamento de Agronomia, Universidade Federal Rural de Pernambuco - UFRPE, Recife, Brazil.
authors.forename: Wellington M
authors.initials: WM
authors.lastname: Silva
authors.affiliation: Departamento de Agronomia, Universidade Federal Rural de Pernambuco - UFRPE, Recife, Brazil.
authors.forename: Ηerbert A A
authors.initials: ΗAA
authors.lastname: Siqueira
authors.affiliation: Rothamsted Research, Harpenden, Herts, UK.
authors.forename: Sofia
authors.initials: S
authors.lastname: Iqbal
authors.affiliation: Rothamsted Research, Harpenden, Herts, UK.
authors.forename: Bartlomiej J
authors.initials: BJ
authors.lastname: Troczka
authors.affiliation: Rothamsted Research, Harpenden, Herts, UK.
authors.forename: Martin S
authors.initials: MS
authors.lastname: Williamson
authors.affiliation: College of Life and Environmental Sciences, University of Exeter, Penryn, UK.
authors.forename: Chris
authors.initials: C
authors.lastname: Bass
authors.affiliation: Hellenic Agricultural Organisation - 'Demeter', Institute of Olive Tree, Subtropical Crops and Viticulture, Heraklion, Crete, Greece.
authors.forename: Anastasia
authors.initials: A
authors.lastname: Tsagkarakou
authors.affiliation: Institute of Molecular Biology & Biotechnology, Foundation for Research & Technology Hellas, Crete, Greece; Department of Crop Science, Agricultural University of Athens, Athens, Greece.
authors.forename: John
authors.initials: J
authors.lastname: Vontas
authors.affiliation: Bayer CropScience, R&D Pest Control, Monheim, Germany. Electronic address: ralf.nauen@bayer.com.
authors.forename: Ralf
authors.initials: R
authors.lastname: Nauen
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001549:Benzamides / Q000494:pharmacology; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics / Q000378:metabolism; D019476:Insect Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D007305:Insecticide Resistance / Q000235:genetics*; D007306:Insecticides; D009036:Moths / Q000235:genetics / Q000502:physiology*; D017354:Point Mutation; D019837:Ryanodine Receptor Calcium Release Channel / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D013450:Sulfones / Q000494:pharmacology; D062367:ortho-Aminobenzoates / Q000494:pharmacology; D005466:Fluorocarbons; D010797:Phthalimides
chemical_list: D001549:Benzamides; D018076:DNA, Complementary; D019476:Insect Proteins; D007306:Insecticides; D019837:Ryanodine Receptor Calcium Release Channel; D013450:Sulfones; D062367:ortho-Aminobenzoates; C517733:chlorantranilipole; C508762:flubendiamide; D005466:Fluorocarbons; D010797:Phthalimides
keywords: Chlorantraniliprole; Diamide resistance; Flubendiamide; Ryanodine receptor; Target-site mutation; Tuta absoluta