Spider mites are one of major pests in cultivations of various ornamental plants and also important in plant quarantine service. Due to the very small body size and high similarity within the Genus the identification of species is difficult even at the microscopic observation. To identify 5 major species (Tetranychus cinnabarinus, T. urticae, T. phaselus, T. kanzawai and T. truncatus) in the Genus Tetranychus at the molecular level, we designed 4 species-specific primer sets using nucleotide sequences of the internal transcribed spacer (ITS2) region in the nuclear ribosomal repeat unit. At the PCR diagnosis of extracted genomic DNAs of 5 species using each primer set, specific primers of both T. phaselus and T. truncates were species-specific to their own species samples. However, specific primer set of T. urticae detected T. cinnabarinus as well as T. urticae. Specific primer set of T. kanzawai detected T. truncates as well as T. kanzawai, even though detection intensities were lower in non-target species.

Due to the very small body size of spider mite, it is difficult to prepare DNA extraction simultaneously with slide samples from a single individual. Here we developed non-mashed DNA extraction method from a single spider mite to apply for molecular as well as morphological identification. Total genomic DNA was isolated from a single female adult using Genomic DNA extraction kit without the sample homogenization. DNA content of a single spider mite was 60-90 ng, which is sufficient for the PCR analysis. However, the quantity of extracted DNA and quality of the cuticle sample were dependent on the incubation time into the lysis buffer. Our results suggest that non-mashed DNA extraction method would be useful for the identification of very small mites as well as insects at the levels of DNA and morphology.

Sweet potato whitefly Bemisia tabaci is a serious pest of various economically important crops. For the control of B. tabaci in an environment-friendly way, we demonstrated the effect of azadirachtin, which is an active conpound of neem oil as an botanical insecticide, on the development of B. tabaci by using an assay of single tomato leaf. Egg hatch rates were 53.6, 50.3% at 5 and 10 ppm, respectively. Adult eclosion rates were 30.0, 22.9% at these doses. We determined the whitefly control efficacies of two application methods of neem-based products by comparing between a direct spray of liquid-type into leaves and a soil treatment of pellet-type. Soil treatment of neem was greatly inhibited adult colonization by 75%. Those plants also inhibited the rates of oviposition and larval development. However, single treatment of foliar spray of neem (5-10 ppm) did not significantly inhibit the initial colonization of adult whiteflies. Furthermore, adult colonization was gradually increased regardless of neem spray.