Seroepidemiology of Toxoplasma gondii and Neospora caninum in seals around Hokkaido, Japan

Serological analysis was performed to detect Toxoplasma gondii and Neospora caninum infection in seals in Hokkaido. Serum samples were collected from 322 Kuril harbor seals (Phoca vitulina stejnegeri) at Nosappu, Akkeshi and Erimo, from 46 spotted seals (P. largha) at Nosappu, Erimo, Yagishiri Island, Hamamasu and Syakotan, and from 4 ribbon seals (P. fasciata) and a bearded seal (Erignathus barbatus) at Nosappu between 1998 and 2006. Recombinant surface antigen of T. gondii (SAG2t) and N. caninum (NcSAG1t) were used as antigens for ELISA to detect antibodies. Antibodies against SAG2t were detected from 4% of 77 Kuril harbor seals at Nosappu in 2005. Antibodies against NcSAG1t were detected from 2% (1/66) in 2003, 5% (4/79) in 2004 and 10% (8/77) in 2005 of Kuril harbor seals and 11% of 9 spotted seals in 2004 sampled at Nosappu. Eight percent of 12 Kuril harbor seals from Akkeshi and 25% of 4 spotted seals from Erimo in 2005 also contained antibodies against NcSAG1t. These suggest sporadic infection of T. gondii and N. caninum in Kuril harbor seals and spotted seals in Hokkaido. Of the ELISA-positive seals, 2 seals having anti-SAG2t antibodies and 3 seals having anti-NcSAG1t antibodies in 2005 were judged to be juveniles that have no maternal antibodies. These suggest that the protozoan infections have occurred in recent years. Infection of terrestrial protozoa such as T. gondii and N. caninum in seals indicates that the sea environment has been contaminated with protozoa.     

Secomycalolide A: A New Proteasome Inhibitor Isolated from a Marine Sponge of the Genus Mycale

A new oxazole-containing proteasome inhibitor, secomycalolide A, together with known mycalolide A and 30-hydroxymycalolide A, was isolated from a marine sponge of the genus Mycale. They showed proteasome inhibitory activities with IC₅₀ values of 11–45 μg/mL.     

Systems for making NIAS Core Collections, single-seed-derived germplasm, and plant photo images available to the research community

The National Institute of Agrobiological Sciences (NIAS) Genebank coordinates the conservation of plant, microorganism, and animal genetic resources related to food and agriculture in Japan. It also coordinates the distribution of genetic resources in the public domain for research, breeding, and educational purposes. To operate the NIAS Genebank efficiently, we have developed a genetic resources database, data management software, and web-based data retrieval systems to make the data available worldwide. This article describes the NIAS Genebank’s Core Collections of global and Japanese soybean (Glycine max (L.) Merr.), Japanese azuki bean (Vigna angularis (Willd.) Ohwi et Ohashi), and Japanese wheat (Triticum aestivum L. s. l.), all of which are available through the Genebank website. This article also describes new features of the NIAS Genebank database, such as the ability to select single-seed-derived germplasm of soybean in the plant search system and to download photographic data on accessions. By using the downloaded plant image PDF files, users can obtain detailed passport and agronomic information by clicking on the image of an accession of interest.     

Development of PCR-based DNA marker for detection of white carrot contamination caused by Y2 locus

In carrot (Daucus carota L.), the taproot colors orange, yellow and white are determined mostly by the Y, Y2, and Or loci. One of the most severe issues in carrot seed production is contamination by wild white carrot. To evaluate the contamination ratio, easily detectable DNA markers for white carrot are desired. To develop PCR-based DNA markers for the Y2 locus, we have re-sequenced two orange-colored carrot cultivars at our company (Fujii Seed, Japan), as well as six white- and one light-orange-colored carrots that contaminated our seed products. Within the candidate region previously reported for the Y2 locus, only one DNA marker, Y2_7, clearly distinguished white carrots from orange ones in the re-sequenced samples. The Y2_7 marker was further examined in 12 of the most popular hybrid orange cultivars in Japan, as well as ‘Nantes’ and ‘Chantenay Red Cored 2’. The Y2_7 marker showed that all of the orange cultivars examined had the orange allele except for ‘Beta-441’. False white was detected in the orange-colored ‘Beta-441’. The Y2_7 marker detected white root carrot contamination in an old open-pollinated Japanese cultivar, ‘Nakamura Senkou Futo’. This marker would be a useful tool in a carrot seed quality control for some cultivars.     

Domain composition of rhamnose-binding lectin from shishamo smelt eggs and its carbohydrate-binding profiles

Osmerus (Spirinchus) lanceolatus egg lectin (OLL) is a member of the rhamnose-binding lectin (RBL) family which is mainly found in aqueous beings. cDNA of OLL was cloned, and its genomic architecture was revealed. The deduced amino acid (aa) sequence indicated that OLL was composed of 213 aa including 95 aa of domain N and 97 aa of domain C. N and C showed 73 % sequence identity and contained both -ANYGR- and -DPC-KYL-peptide motifs which are conserved in most of the RBL carbohydrate recognition domains. The calculated molecular mass of mature OLL was 20,852, consistent with the result, and 20,677.716, from mass spectrometry. OLL was encoded by eight exons: exons 1 and 2 for a signal peptide; exons 3–5 and 6–8 for N- and C-domains, respectively. Surface plasmon resonance spectrometric analyses revealed that OLL showed comparable affinity for Galα- and β-linkages, whereas Silurus asotus lectin (SAL), a catfish RBL, bound preferentially to α-linkages of neoglycoproteins. The Kd values of OLL and SAL against globotriaosylceramide (Gb3) were 1.69 × 10^?5 M for and 2.81 × 10^?6 M, respectively. Thus, the carbohydrate recognition property of OLL is slightly different from that of SAL. On the other hand, frontal affinity chromatography revealed that both OLL and SAL interacted with only glycolipid-type oligosaccharides such as Gb3 trisaccharides, not with N-linked oligosaccharides. The domain composition of these RBLs and an analytical environment such as the “cluster effect” of a ligand might influence the binding between RBL and sugar chains.     

Overexpression of the LASAP2 gene for secretory acid phosphatase in white lupin improves the phosphorus uptake and growth of tobacco plants

Secretion of acid phosphatase (APase) from the roots to take up phosphorus (P) is a well-known strategy of plants under P-deficient conditions. White lupin, which shows vigorous growth in low-P soils, is noted for its ability to secrete APase under P-deficient conditions. The APase secreted by white lupin roots is stable in soil solution and shows low substrate specificity, suggesting that genetic modification of plants using the APase gene LASAP2 might improve their ability to use organic P. The objective of the present study was to evaluate the potential of LASAP2 transgenic plants to increase organic P utilization. Dry matter production and P accumulation were higher in LASAP2 transgenic tobacco plants grown in gel media containing soluble phytate as the sole P source than in wild-type tobacco plants. Phosphorus uptake by the transgenic plants also increased in soil culture conditions. LASAP2 was apparently more effective in the liberation of organic P, including phytate, in the soil than the native tobacco APase. Thus, the enzymatic stability of LASAP2 in the soil appears to be an important factor for P acquisition.     

Development of EST-SSR markers and construction of a linkage map in faba bean (Vicia faba)

To develop a high density linkage map in faba bean, a total of 1,363 FBES (Faba bean expressed sequence tag [EST]-derived simple sequence repeat [SSR]) markers were designed based on 5,090 non-redundant ESTs developed in this study. A total of 109 plants of a ‘Nubaria 2’ × ‘Misr 3’ F₂ mapping population were used for map construction. Because the parents were not pure homozygous lines, the 109 F₂ plants were divided into three subpopulations according to the original F₁ plants. Linkage groups (LGs) generated in each subpopulation were integrated by commonly mapped markers. The integrated ‘Nubaria 2’ × ‘Misr 3’ map consisted of six LGs, representing a total length of 684.7 cM, with 552 loci. Of the mapped loci, 47% were generated from multi-loci diagnostic (MLD) markers. Alignment of homologous sequence pairs along each linkage group revealed obvious syntenic relationships between LGs in faba bean and the genomes of two model legumes, Lotus japonicus and Medicago truncatula. In a polymorphic analysis with ten Egyptian faba bean varieties, 78.9% (384/487) of the FBES markers showed polymorphisms. Along with the EST-SSR markers, the dense map developed in this study is expected to accelerate marker assisted breeding in faba bean.     

Evaluation of site-specific management zones on a farm with 124 contiguous small paddy fields in a multiple-cropping system

The identification of homogeneous management zones (MZs) within a field is a basis for site-specific management (SSM). We assessed the method of defining MZs based on the spatio-temporal homogeneity of six soil properties and above-ground biomass data from paddy rice, winter wheat and soybean over 3 years on a farm with 124 contiguous small paddy fields. The soil data were recorded at 372 soil sampling sites on a rectangular grid over the farm. A non-hierarchical cluster analysis was applied to the soil data and the algorithm grouped the sites into three clusters with similar soil properties. These clusters represent soil fertility and soil drainage. The three clusters were not randomly distributed across the fields, but formed contiguous areas associated with landscape position. This was due to the spatial variation of the soil in the study area. We delineated five MZs based on the spatial structure of the soil heterogeneity of the study area. The validity of the MZs was evaluated using the biomass data from paddy rice, winter wheat and soybean in each MZ; this depended mainly on soil fertility when conditions were dry. When the growing season precipitation was greater than the 10-year average, the biomass of winter wheat and soybean depended on soil drainage. This suggested that the delineation of MZs for site-specific management in fields under a paddy-upland crop rotation system should be based on several soil properties. The biomass data from the three crops over 3 years was not effective for delimiting MZs.     

Chromosome doubling of Lychnis spp. by in vitro spindle toxin treatment of nodal segments

Lychnis (Caryophyllaceae) consists of about 30 species distributed throughout the temperate regions of the Northern Hemisphere, from East Asia to Europe. Many Lychnis spp. have high ornamental value and cultivated as pot or garden plants. In the present study, in vitro chromosome doubling of several Lychnis spp. was examined in order to widen their variability in horticultural traits. Initially effect of various spindle toxin treatments [100, 500 or 1000 mg l⁻¹ colchicine (COL), 10, 20 or 50 mg l⁻¹ oryzalin (ORY), or 1, 5, 10 mg l⁻¹ amiprophos-methyl (APM)] of nodal segments of a triploid genotype of L. senno (3x) was investigated on survival of nodal segments and chromosome doubling in nodal segment-derived plantlets. Significantly higher percentage (75.0%) of surviving segments after spindle toxin treatment was obtained in 10 mg l⁻¹ ORY treatment. Flow cytometry (FCM) analysis of leaf tissues showed that 9.4–13.8% of plantlets, which were derived from 10 to 20 mg l⁻¹ ORY, or 5 mg l⁻¹ APM treatments, were hexaploid (6x) or ploidy chimera (3x + 6x, 4x + 6x, 5x + 6x, 3x + 4x + 6x). The results obtained by FCM analysis were confirmed by chromosome observation in root tip cells. Thus 10 mg l⁻¹ ORY treatment of nodal segments is suitable for in vitro chromosome doubling of triploid L. senno. Efficient chromosome doubling was also achieved in diploid L. fulgens (2x) and L. sieboldii (2x) by treating nodal segments with 10 mg l⁻¹ ORY: 68.9–88.7% of nodal segments survived after ORY treatment, and 24.7–26.5% of plantlets derived from ORY-treated nodal segments were tetraploid (4x) or ploidy chimera (2x + 4x) in both species. These results indicate that the in vitro chromosome doubling method established for triploid L. senno may be applicable to a wide range of Lychnis spp. Tetraploid L. fulgens and L. sieboldii showed a compact plant form, and had thick stems and deep green leaves compared with the diploid mother plants. On the other hand, hexaploid L. senno showed very poor growth and died before flowering.     

Airborne basidiospores as an inoculum source of <Emphasis Type="Italic">Typhula variabilis</Emphasis> and the effect of hilling on the incidence of Typhula winter rot of carrots

Typhula winter rot on overwintering carrots caused by Typhula variabilis is a newly confirmed disease, and no practical control measure is yet available. To develop a control method, here we researched the infection period of T. variabilis and the time that winter rot appeared on carrots. Using spore traps, we found that basidiospore rain occurred from September to November before snowfall in Memuro, Hokkaido. In addition, carrot leaves collected in autumn had already been infected by T. variabilis. These epidemiological investigations revealed that the pathogen releases basidiospores to infect carrot leaves before snow cover, resulting in root decay under snow. An effective control method was then developed to avoid direct contact of basidiospores of T. variabilis with plant tops by covering the plants with soil in autumn. Thus, the percentage of rotted roots was reduced to about half.