Use of bioassays and molecular markers to detect insecticide resistance in stored products beetles

The qualitative and quantitative losses caused by stored product insects are of great concern, and since there is only a few active ingredients available for their control it is very important to have a frequent insect resistance monitoring. The objective of this research is to evaluate combination of bioassays and molecular marker techniques to detect insecticide resistance in stored product beetles. The Coleoptera species used for the tests were Sitophilus oryzae (L.) (Curculionidae), Rhyzopertha dominica (F.) (Bostrichidae) and Oryzaephilus surinamensis (L.) (Silvanidae). For the bioassays it was used the impregnated filter paper technique, applying 1 mL of deltamethrin (K-Obiol 25 CE) using four concentrations and five replicates, including a control with solvent only. Ten adults of each species were liberated separately on each dish. The mortality was evaluated after 24 h and resistance determined by probit analysis. The samples used for the PCR-RAPD were either in vivo or preserved in 70% ethanol, kept in -18 degree C freezer. After extraction, quantification and DNA quality analysis, the 25 mu L samples had the DNA amplified and tested with six primers. The bioassays showed a crescent mortality proportional to insecticide concentration. The resistance factor for R. dominica, S. zeamais and S. oryzae were: 2,2; 3,2 and 9,2, respectively, compared to the susceptible populations of each species. The PCR-RAPD analysis revealed bands which indicate inter and intraspecific variability in the populations, but it was not possible to correlate them to resistance. The association of bioassay and PCR-RAPD represents a precise and valuable tool for resistance management of stored product insects, but more populations and primers should be tested.