title: Phosphine resistance in Australian Cryptolestes species (Coleoptera: Laemophloeidae): perspectives from mitochondrial DNA cytochrome oxidase I analysis.
abstract: BACKGROUND
  The flat grain beetle (FGB) species Cryptolestes ferrugineus, C. pusillus, C. pusilloides and C. turcicus are major stored-product pests worldwide, of which the first three are present in Australia. C. ferrugineus is also a species with high phosphine resistance status in various countries. Morphological identification of Cryptolestes species is difficult and represents an additional barrier to effective management of phosphine resistance in FGBs.
  
  
  RESULTS
  Mitochondrial DNA cytochrome oxidase I (mtDNA COI) gene characterisation enabled differentiation of the four major FGB pest species through direct sequence comparison, and enabled the development of a PCR-RFLP method for rapid species differentiation. We detected two mtDNA haplotypes (Cunk-01, 02) present at low frequencies with an average nucleotide divergence rate of 0.079 ± 0.011 (SE) from C. pusillus. This nucleotide divergence rate is similar to that between C. ferrugineus and C. pusilloides (0.088 ± 0.012). Male and female genitalia morphologies of the Cunk-02 individuals indicated they were consistent with C. pusillus, yet DNA sequence analyses suggested species-level divergence. The mtDNA COI gene of phosphine-bioassayed, lab-reared F1 generation survivors supported the presence of strong phosphine resistance in C. ferrugineus, but unexpectedly also in C. pusilloides and C. pusillus F1 survivors.
  
  
  CONCLUSIONS
  We demonstrated the utility of molecular DNA techniques for differentiating closely related insect species, and its usefulness in assisting the management of pest insect species. The likely presence of a cryptic C. pusillus species in Australia and the possible development of strong phosphine resistance in Australian FGB pest species require further investigation. © 2014 Society of Chemical Industry.
authors.affiliation: Biosecurity Flagship, CSIRO Ecosystem Sciences, Black Mountain Laboratories, ACT, Australia.|Plant Biosecurity Cooperative Research Centre, Bruce, ACT, Australia.
authors.forename: Wee Tek
authors.identifier: http://orcid.org/0000-0002-8451-0811
authors.initials: WT
authors.lastname: Tay
authors.affiliation: Biosecurity Flagship, CSIRO Ecosystem Sciences, Black Mountain Laboratories, ACT, Australia.
authors.forename: Stephen J
authors.initials: SJ
authors.lastname: Beckett
authors.affiliation: CSIRO Ecosystem Sciences, Brisbane, Australia.
authors.forename: Paul J
authors.initials: PJ
authors.lastname: De Barro
mesh_terms: D000818:Animals; D001315:Australia; D001681:Biological Assay / Q000379:methods; D001517:Coleoptera / Q000187:drug effects / Q000201:enzymology; D004272:DNA, Mitochondrial / Q000235:genetics*; D003576:Electron Transport Complex IV / Q000235:genetics*; D005260:Female; D019476:Insect Proteins / Q000235:genetics; D007305:Insecticide Resistance / Q000235:genetics; D007306:Insecticides / Q000494:pharmacology*; D008297:Male; D010720:Phosphines / Q000494:pharmacology*; D017422:Sequence Analysis, DNA
chemical_list: D004272:DNA, Mitochondrial; D019476:Insect Proteins; D007306:Insecticides; D010720:Phosphines; D003576:Electron Transport Complex IV; C044646:phosphine
keywords: Cryptolestes; biosecurity; cryptic species; mtDNA COI; phosphine; stored grain