Phosphine resistance in Australian Cryptolestes species (Coleoptera: Laemophloeidae): perspectives from mitochondrial DNA cytochrome oxidase I analysis

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 . RESULT 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 F 1 generation survivors supported the presence of strong phosphine resistance in C. ferrugineus , but unexpectedly also in C. pusilloides and C. pusillus F 1 survivors. CONCLUSION 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