Construction of genetic linkage map and identification of a novel major locus for resistance to pine wood nematode in Japanese black pine (Pinus thunbergii).

BACKGROUND: Pine wilt disease (PWD), which is caused by the pine wood nematode (PWN) Bursaphelenchus xylophilus, is currently the greatest threat to pine forests in Europe and East Asian countries including Japan. Constructing a detailed linkage map of DNA markers and identifying PWD resistance genes/loci lead to improved resistance in Pinus thunbergii, as well as other Pinus species that are also susceptible to PWD. RESULTS: A total F1 mapping population of 188 individuals derived from a cross between the PWD-resistant P. thunbergii varieties 'Tanabe 54' (resistant rank 2 to PWD) and 'Tosashimizu 63' (resistant rank 4 to PWD) was inoculated with PWN, and was evaluated for disease symptoms. To perform linkage analysis for PWN resistance, a set of three maps was constructed; two parental maps generated using the integrated two-way pseudo-testcross method, and a consensus map with population-type cross-pollination. The linkage map of 'Tanabe 54' consisted of 167 loci, and covered 14 linkage groups (LGs), with a total genetic distance of 1214.6 cM. The linkage map of 'Tosashimizu 63' consisted of 252 loci, and covered 14 LGs, with a total genetic distance of 1422.1 cM. The integrated consensus map comprised 12 LGs with the basic chromosome number of P. thunbergii, and a total genetic distance of 1403.6 cM. Results from quantitative trait loci (QTL) analysis using phenotype data and linkage maps indicated that PWN resistance is controlled by a single dominant allele, which was derived from the 'Tanabe 54' female parent. This major QTL was located on linkage group 3 and was designated PWD1 for PINE WILT DISEASE 1. CONCLUSIONS: The PWD1 locus is a major resistance QTL located on the Pinus consensus LG03 that acts in a dominant manner to confer pine wood nematode resistance. Information from the present study will be useful for P. thunbergii breeding programs to improve resistance to PWD, and also to help identify susceptibility genes in Pinus species.

Correction: Evaluation of thermoregulation of different pine organs in early spring and estimation of heat reward for the western conifer seed bug (Leptoglossus occidentalis) on male cones.

[This corrects the article DOI: 10.1371/journal.pone.0272565.].

A multi-ring optical packet and circuit integrated network with optical buffering.

We newly developed a 3 × 3 integrated optical packet and circuit switch-node. Optical buffers and burst-mode erbium-doped fiber amplifiers with the gain flatness are installed in the 3 × 3 switch-node. The optical buffer can prevent packet collisions and decrease packet loss. We constructed a multi-ring optical packet and circuit integrated network testbed connecting two single-ring networks and a client network by the 3 × 3 switch-node. For the first time, we demonstrated 244 km fiber transmission and 5-node hopping of multiplexed 14-wavelength 10 Gbps optical paths and 100 Gbps optical packets encapsulating 10 Gigabit Ethernet frames on the testbed. Error-free (frame error rate < 1 × 10(-4)) operation was achieved with optical packets of various packet lengths. In addition, successful avoidance of packet collisions by optical buffers was confirmed.

Quantitative Trait Loci Analysis Based on High-Density Mapping of Single-Nucleotide Polymorphisms by Genotyping-by-Sequencing Against Pine Wilt Disease in Japanese Black Pine (Pinus thunbergii).

Identifying genes/loci for resistance to pine wilt disease (PWD) caused by the pine wood nematode (PWN) is beneficial for improving resistance breeding in Pinus thunbergii, but to date, genetic information using molecular markers has been limited. Here, we constructed a high-density linkage map using genotyping-by-sequencing (GBS) and conducted quantitative trait loci (QTL) analysis for PWD resistance for the self-pollinated progeny of "Namikata 73," which is the most resistant variety among resistant varieties of P. thunbergii, following inoculation tests with PWN. An S1 mapping population consisting of the 116 progenies derived from self-pollination of the resistant variety, "Namikata 73" (resistance rank 5 to PWN), was inoculated with PWN isolate Ka-4 and evaluated for disease symptoms. To construct a high-density linkage map, we used single-nucleotide polymorphisms (SNPs) identified by GBS based on next-generation sequencing technology and some anchor DNA markers, expressed sequence tag (EST)-derived SNP markers and EST-derived simple sequence repeat (SSR) markers, and genomic SSR markers. The linkage map had 13 linkage groups (LGs) consisting of 2,365 markers including 2,243 GBS-SNP markers over a total map distance of 1968.4 centimorgans (cM). Results from QTL analysis using phenotype data and the linkage map indicated that PWD resistance is controlled by a single locus located on LG-3, as identified in a previous study. This locus showed overdominant genetic action in the present study. With the confirmation of PWD1 in two different mapping populations (present study and a previous study), the locus associated with this region is thought to be a good target for marker-assisted selection in P. thunbergii breeding programs in order to obtain high levels of resistance to PWD caused by PWN.

Ribosomal DNA haplotype distribution of Bursaphelenchus xylophilus in Kyushu and Okinawa islands, Japan.

Ribosomal DNA region sequences (partial 18S, 28S and complete ITS1, 5.8S, and ITS2) of the pinewood nematode (Bursaphelenchus xylophilus) were obtained from DNA extracted directly from wood pieces collected from wilted pine trees throughout the Kyushu and Okinawa islands, Japan. Either a 2569bp or 2573bp sequence was obtained from 88 of 143 samples. Together with the 45 rDNA sequences of pinewood nematode isolates previously reported, there were eight single nucleotide polymorphisms and two indels of two bases. Based on these mutations, nine haplotypes were estimated. The haplotype frequencies differed among regions in Kyushu island (northwest, northeast and center, southeast, and southwest), and the distribution was consistent with the invasion and spreading routes of the pinewood nematode previously estimated from past records of pine wilt and wood importation. There was no significant difference in haplotype frequencies among the collection sites on Okinawa island.

Development of a simple analytical method to determine arsenite using a DNA aptamer and gold nanoparticles.

A simple analytical method was developed to determine the arsenite (As(III)) concentration using a DNA aptamer and gold nanoparticles (AuNPs). Prior to sample measurements, the method sensing mechanism was confirmed by analyzing the particle size of the AuNPs at each step of the analysis procedure, and the key operational parameters that affect the method performance were optimized. The optimal final NaCl concentration, incubation time with NaCl and pH of a 3-(N-morpholino) propanesulfonic acid buffer were 60 mM, 10 min and 7.3, respectively. A calibration curve was created under optimized operational conditions. The calibration curve was linear from a 1.0- to 10-µM As(III) concentration. The detection limit was 2.1 µM (161 µg/L). Using the calibration curve, we evaluated groundwater samples spiked with As(III). As(III) concentrations in groundwater pretreated with a 0.2-µm-pore-size membrane filter and cation-exchange resin were determined by using the method, which suggests that the proposed method can be used to determine the As(III) concentration in groundwater.