DNA-based identification of spider mites: molecular evidence for cryptic species of the genus Tetranychus (Acari: Tetranychidae).

Spider mites are difficult to identify because they are very small and have a limited number of diagnostic characters. Most species of the spider mite genus Tetranychus in Japan are morphologically similar, differing only in the diameter of the aedeagal knob in males. Because this genus contains many important pests, the unambiguous identification of species is crucial for effective pest management and quarantine procedures. DNA-based methods could complement the morphological methods. We examined whether Tetranychus species in Japan could be identified by DNA sequences using the internal transcribed spacer region of nuclear ribosomal DNA and the cytochrome c oxidase subunit I gene of mitochondrial DNA. We determined sequences of the 13 known Tetranychus species in Japan. We could identify 10 of the 13 species in the internal transcribed spacer tree. In the cytochrome c oxidase subunit I tree, we could identify all 13 known Tetranychus species in Japan. Although Tetranychus kanzawai Kishida and T. parakanzawai Ehara were identified by DNA sequences, they were clearly separated into two monophyletic clades each, indicating that a cryptic species existed in each species.

Molecular-based identification and phylogeny of Oligonychus species (Acari: Tetranychidae).

The genus Oligonychus has been morphologically divided into two groups based on the direction of curvature of the aedeagus and includes some morphologically similar species that are difficult to distinguish. To develop DNA-based methods for identifying Oligonychus species and to determine the phylogenetic relationships among them, we examined the cytochrome c oxidase subunit I gene of mitochondrial DNA and the internal transcribed spacer and 28S regions of nuclear ribosomal RNA gene for 17 species. Based on the genetic distances (p-distances) of the three DNA regions, the range of intraspecific divergence was found to be below (and not overlap) the range of interspecific divergence, which allowed the 17 species to be discriminated correctly, consistent with their classification based on morphology. Phylogenetic trees constructed by neighbor-joining and Bayesian methods clearly showed two clades, consisting of species whose aedeagi curve ventrally and dorsally, respectively. Three Oligonychus species inhabiting gramineous plants formed clearly defined subclades.

Isolation and characterization of polymorphic microsatellite loci from the water mite Hygrobates fluviatilis (Acari: Hydrachnidia: Hygrobatidae).

We developed and characterized 13 microsatellite markers from the water mite Hygrobates fluviatilis. The genotypes of 32 diploid females were assessed, and all of the loci were polymorphic. The number of alleles per locus ranged from two to 21, and the observed heterozygosity ranged from 0.063 to 0.813. These microsatellite markers are the first published for water mites and will contribute to research on the population structure of this widespread species.

Decreased risk of catheter infection in infants and children using subcutaneous tunneling for continuous caudal anesthesia.

Continuous caudal anesthesia has been commonly used for intra- and post-operative analgesia in infants and children. However, it has a potential risk of bacterial infection, especially in infants in whom the catheter site is easily contaminated with loose stool. To avoid infection, the authors applied a new procedure using subcutaneous tunneling for continuous caudal anesthesia. In the 18 cases studied with subcutaneous tunneling, clinical signs of infection were absent and bacterial colonization was not found on the catheter tip after 3.9 +/- 1.4 days of catheterization. The incidence of catheter colonization after continuous caudal anesthesia without tunneling had been reported. In their reports, the incidence of catheter colonization ranged from 20% to 37%. Therefore, caudal catheterization with subcutaneous tunneling is a simple and safe method, and has proved very effective to reduce the risk of epidural infection.

Genetic diversity and phylogeny of the Kanzawa spider mite, Tetranychus kanzawai, in Japan.

Two types are known in the Kanzawa spider mite, Tetranychus kanzawai (K and T; see Gotoh et al., 1999), which differ in host range and have a unidirectional incompatibility. Prior to DNA analyzes, crossing between females of a known K type and males of each of 17 strains collected in Japan showed that six of the strains were of the K type, live were the T type and the rest consisted of a mixture of the two types. In order to elucidate the genetic diversity and phylogenetic relationship of T kanzawai in Japan, we analyzed the DNA sequences of two regions--the internal transcribed spacer 1 (ITS1) of the nuclear ribosomal DNA (rDNA) and a fragment of the cytochrome oxidase subunit I gene (COI) of mitochondrial DNA - using 11 strains (six K-type strains and five T-type strains). Base substitutions were detected on 25 sites of COI (375bp) and 19 sites of ITS1 (486bp), resulting in eight and 17 haplotypes, respectively. The phylogenetic trees constructed using the DNA sequences failed to clearly distinguish between the two types. The results suggested that the T type was derived from the K type.

Molecular cloning and characterization of a microsatellite locus found in an RAPD marker of a spider mite, Panonychus citri (Acari: Tetranychidae).

Genetic markers were searched using PCR with 40 kinds of decanucleotide primers to investigate DNA polymorphism in Panonychus citri. A region consisting of a variable number of CT tandem repeats (microsatellite) was found in a fragment amplified with the OPB10 primer. The microsatellite differed in size by ca. 100bp among several P. citri populations screened and was derived from at least seven alleles. This region was characteristic of P. mori and P. osmanthi, but was lacking in P. ulmi. The flanking regions were highly conserved among these species.