Molecular techniques for the detection of granary weevil (<i>Sitophilus granarius</i>L.) in wheat and flour

Abstract

Abstract\n Abstract &#13;\nThe granary weevil (Sitophilus granarius L.) is a stored grain pest which causes major economic losses. It reduces the quantity and quality of the grain by its feeding and excretion.&#13;\nWe analyzed sequences of S. granarius mitochondrial cytochrome oxidase subunits genes: mtCOI and mtCOII and compared them with mtCOI/II sequences available in GenBank. The analyzed genes displayed a high level of homology between corresponding subunits. We have undertaken attempts to develop detection methods of contamination by S. granarius in wheat and wheat flour based on the molecular biology techniques: standard and real-time PCR with a TaqMan molecular probe. We applied specific primers designed on the basis of available sequences for mtCOI and mtCOII genes, and established optimal conditions of reactions. The specificity of both methods was studied by using a species closely related to S. granarius ? S. oryzae and S. zeamais. We showed that the sensitivity threshold was very high ? we were able to detect the equivalent of one beetle per 100 kg of flour when the real-time PCR with TaqMan probe method was applied to model samples. The primer sets used turned out to be species specific, and technique was rapid, reliable and very sensitive.