Insect DNA Exposed to Two Insecticides

• Analyses of DNA is now a standard method in exploring taxonomical relationships between taxa or when studying intraspecific genetic variation. For these purposes museum specimen are often used. During their “museum history”, biological collections often have been treated with various insecticides to avoid museum pest attacks. There is a risk that some insecticides have more or less destroyed the specimen DNA, and thus it is important to know to what extent this happens to be able to estimate if old collection are useful for DNA screening and also to avoid future mistakes. Kigawa et al. (2003) showed that methyl bromide, methyl bromide/ethylene oxide (mixed), ethylene oxide, propylene oxide and methyl iodide all caused degradation of DNA in samples of mushroom and chicken muscle. We have tested possible negative effects on insect DNA from exposure of paradichlorobenzene and diclorvos. Species used were Schistocerca gregaria (Orthoptera), Musca domestica (Diptera), Dermestes hemeroidalis (Coleoptera), Periplaneta americana (Blattodea). The specimens were cut into two parts before completely dried in silica gel in closed containers for about 2 weeks. Three serials were set up in sealed, small glass containers for each species: (1) paradichlorobenzene (0.5g ±0.02g, representing saturated concentration); (2) diclorvos (0.5g ±0.02g, representing saturated concentration); and (3) no chemicals (blank). Each serial was sampled after 1 and 4 weeks. Immediately after sampling, the insect tissues were exposed to clean air for 8 hours in order to eliminate chemicals from the tissue before subsequent treatment. Together with the blank samples, the exposed tissues were stored in deep freezers before extracted for DNA after the last tissue sampling. After extraction, a 658 bp (base pair) COI gene fragment of the DNA was amplified using the forward and reverse primers HCO2198 (TAA ACT TCA GGG TGA CCA AAA AAT CA) and HCOout (CCA GGT AAA ATT AAA ATA TAA ACT TC), respectively. Electrophoresis of the PCR products was run for about 2h and presence or absence of the amplified COI gene fragment showed as band or absence of band in UV light. The gel was photographed for documentation of the results. For S. gregaria we did not obtain any clear results, probably because the primer did not work for the COI gene of this species. For the other three species no effect could be seen from the exposure of paradichlorobenzene, while dichlorvos destroyed the COI gene fragment after 28 days. For M. domestica a band was present after one week exposure. Our conclusion is that dichlorvos has a deteriorating effect on DNA. A more extensive experimental series on M. domestica has been started at the Swedish Museum of Natural History. Reference: Kigawa, R., Nochide, H., Kimura, H. and Miura, S., Effects of various fumigants, thermal methods and carbon dioxide treatment on DNA extraction and amplification: A case study on freeze-dried mushroom and free-dried muscle specimens. Collection Forum 2003, 18 (1-2): 74-89.

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