We provide the first report on Stigmaeopsis miscanthi Saito, which was identified from Chinese silver grass Miscanthus sinensis on Ulleung Island in Korea. This species is one of the S. celarius complex, which involves several cryptic species. S. miscanthi has longer second dorsal propodosomal setae (P2), which is an important characteristic for species identification in the genus Stigmaeopsis. We determined nucleotide sequences of the internal transcribed spacer 2 (ITS2) and cytochrome c oxidase subunit I (COI) of S. miscanthi. The COI sequence differed by 0.6% between Korean and Japanese strains. The comparison between S. miscanthi and S. celarius showed that ITS2 and COI differed by 7.2% and 7.9%, respectively. In addition, species-specific primer sets of both species were designed to show the species classification within the genus.

The genus Panonychus has been reported only two species, P. ulmi and P. citri, in Korea. Two new species, P. mori Yokoyama, 1929 and P. caglei Mellot, 1968 were firstly identified from jujube orchards in Gyeongsan and kudzu vine in Byunsan peninsula in Korea. Morphological differences among four species have been described especially in aedeagus shape. Comparison of nucleotide sequences of both the internal transcribed spacer 2 (ITS2) region of nuclear ribosomal RNA gene and the mitochondrial cytochrome oxidase subunit I (COI) gene were compared between four species. Phylogenetic analysis of ITS2 and COI sequences using neighbor-joining method showed that P. mori and P. caglei were most similar to each other and more closely related to P. ulmi than P. citri. In addition, species-specific primer sets of each species were designed based on ITS2 sequencesand can be used to diagnose species in this genus.

Tetranychid mites are one of the most diverse group including at least 1,200 species in the world. Species identification is difficult due to the small size and similar morphology within the group. We collected 17 species of spider mites from various host plants in different regions of Korea and determined species identity by the comparison of morphological characters and nucleotide sequences of internal transcribed spacer 2 (ITS2) and the cytochrome oxidase subunit I (COI). In addition, we report three new species that were firstly identified in Korea. Amphitetranychus quercivorus (Ehara and Gotoh) was collected from Mongolian oak plant in Daegu, Schizotetranychus miscanthi Saito was from the common reed plant in Ulleungdo, and S. cercidiphylli Ehara was from Bamboo plant in Jeju. Morphological identification of three species were similar with those of Japanese samples, but the ITS2 and COI sequences of A. quercivorus and the COI sequence of S. miscanthi were different with Japanese species at the rates of 1/419, 2/331 and 3/332 nucleotides, repsectively. S. cercidiphylli can be identified by the aedeagus shape of males but we firstly sequenced ITS2 and COI of this species. Our results can be used for the identification of spider mites which are important in plant quarantine.

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.