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.     

Invasion and colonization of mature apple fruit by Erwinia amylovora tagged with bioluminescence genes

Invasion and colonization of mature apple fruit by a transformant of Erwinia amylovora tagged with bioluminescence genes from Vibrio fischeri was examined. The transformant was deposited on cut surfaces of fruit stems, wounds on the shoulders and calyces, injured fruit-bearing twigs of harvested apple fruit, and cut fruit flesh. After incubation in closed stainless steel or plastic boxes at 25°C, fruit were periodically observed with a two-dimensional luminometer. The presence of the transformant in luminous areas was confirmed by isolating it on selective media. E. amylovora, when deposited in fruit stems: (1) can invade mature as well as immature apple fruit; (2) vertically and horizontally spreads and colonizes along vascular bundles, increasing its population; (3) reaches the calyx end and the flesh just under the exocarp within 3–4 days after inoculation; (4) when deposited on cut fruit flesh, irrespective of its maturity, can easily increase its population and survive 2–4 weeks or more at 25°C; and (5) even at the time of fruit maturation, can migrate within twigs rapidly and reaches the abscission layers between fruit-bearing twigs and fruit stems.     

Annual variation of natural 15N abundance in tea leaves and its practicality as an organic tea indicator

To obtain basic knowledge about the relationship between the application of organic fertilizers and the δ15N values of leaves of organically grown tea plants, annual variations in the δ15N values of the tea leaves were investigated. Although variations did not immediately arise after the application of organic fertilizers, differences in the δ15N values between organic and conventional cultivations appeared basically after three years from the beginning of the organic cultivation except when an organic fertilizer with a low δ15N value was applied, and the variation depended on the δ15N values of the fertilizers. In addition, the effectiveness of the δ15N values as a practical indicator of organic teas was examined. The tea leaves collected from organic farms did not always have higher δ15N values than the commercially available nonorganic teas. This result demonstrates that it is not easy to discriminate organic teas from nonorganic teas simply by their δ15N values.     

An in vitro screening assay to discover novel inhibitors of 4-coumarate:CoA ligase

4-Coumarate:CoA ligase (4CL, EC 6.2.1.12) exists only in plants and plays an important role in the phenylpropanoid pathway. Identification of inhibitors targeting 4CL provides a novel approach for developing effective plant growth inhibitors (PGIs). The full-length gene of tobacco 4CL (Nt4CL1) was cloned and expressed in Escherichia coli Cast & Chalm. The recombinant 4CL protein was extracted and purified by several purification steps including gel-filtration and anion-exchange chromatography. 4CL activity assay was miniaturized and optimized using a 96-well microplate and a reader. Among 28 existing herbicides, propanil and swep strongly inhibited in vitro 4CL enzyme activity, and they were selected for further studies. The process of this assay can be developed into a high-throughput screening system of PGI targeting 4CL in the phenylpropanoid pathway.     

Acibenzolar-S-methyl efficacy against bacterial brown stripe caused by Acidovorax avenae subsp. avenae in creeping bentgrass

Journal of General Plant Pathology - Bacterial brown stripe caused by Acidovorax avenae subsp. avenae (Aaa) causes serious damage on creeping bentgrass on golf course greens. However, there are few...     

First report of bacterial blight of crucifers caused by Pseudomonas cannabina pv. alisalensis in Japan

In October 2010, a bacterial disease produced flecks and spots on leaves of Chinese cabbage, cabbage and Japanese radish in Nagano Prefecture, Japan. The symptoms started on the abaxial surface of leaves as angular, water-soaked flecks of 1–2 mm in diameter with a yellow halo of 3–4 mm width. These flecks then became visible on both leaf surfaces, enlarged and coalesced into large blight lesions. The symptoms were similar to bacterial leaf spot caused by Pseudomonas syringae pv. maculicola. The bacterium isolated from leaf lesions formed a white colony and produced polysaccharides on YP agar. The isolates were identified as P. syringae group by LOPAT tests and the 16S rDNA sequence. Moreover, the results of pathogenicity on cruciferous plants, bacteriological characteristics, rep-PCR and the sequences of rpoD and gyrB showed that the isolates should be identified as P. cannabina pv. alisalensis (recently transferred from P. syringae pv. alisalensis). This is the first report of P. cannabina pv. alisalensis isolated from diseased crucifers in Japan.     

Relationship between resistance to pine wilt disease and the migration or proliferation of pine wood nematodes

To study the relationship between resistance to pine wilt disease and the migration or proliferation of pine wood nematodes (PWN) (Bursaphelenchus xylophilus), we conducted experiments using clonally-propagated Japanese black pines (Pinus thunbergii) with pre-evaluated individual resistance levels. Bark including the cortical resin canals—one of the main migration pathways of PWN—was removed by girdling, but neither the migration of PWN nor symptom development of pine wilt disease were inhibited by this treatment in non-resistant clones. Histological observations showed no significant differences in the lumen area or the number of cortical- and xylem- axial resin canals between resistant and susceptible clone groups from a half-sib family. A bioassay using methanol extracts from resistant and susceptible clones showed that extracts from both clones showed similar attractant effects to PWN, but neither had repellent effects. The resistant clones were multi-inoculated with PWN into three split points to mimic migration in the stem. The proportion of damaged plants was not significantly different from that in single-inoculated plants (control). In this experiment, the number of PWN detected from partially-damaged plants was much higher than that from non-damaged plants. An inoculation test using stem cuttings showed that the population of PWN increased in susceptible cuttings at 1–20 days after inoculation (dai), while it remained unchanged or gradually decreased in resistant cuttings. These findings suggested that the factors contributing to resistance were associated with inhibiting the proliferation of PWN, rather than inhibiting their migration.