Reduced sensitivity to fenarimol in Japanese field strains of Venturia nashicola

Monoconidial strains of Venturia nashicola Tanaka et Yamamoto were isolated from Japanese or Chinese white pear trees which had never been treated with sterol demethylation inhibitors (DMIs) and their baseline sensitivities to fenarimol were determined by mycelial growth tests on fungicide-amended culture media. Strains were also obtained from Japanese pear orchards, which had been intensively treated with DMIs for several years and monitored for the shifts of fenarimol sensitivity in comparison with the baseline sensitivity. Results suggested slight shifts to lower fenarimol sensitivity in strains isolated from DMI-treated Japanese pear orchards. However, in inoculation tests on pear seedlings, fenarimol still provided adequate control of V. nashicola strains with reduced sensitivity to fenarimol in vitro, suggesting that the performance of this fungicide will still be maintained in the field. © 1998 Society of Chemical Industry     

Temporal shifts in diversity and quantity of nirS and nirK in a rice paddy field soil

Denitrification is an important part of the nitrogen cycle in the environment, and diverse bacteria, archaea, and fungi are known to have denitrifying ability. Rice paddy field soils have been known to have strong denitrifying activity, but the microbes responsible for denitrification in rice paddy field soils are not well known. Present study analyzed the diversity and quantity of the nitrite reductase genes (nirS and nirK) in a rice paddy field soil, sampled four times in one rice-growing season. Clone library analyses suggested that the denitrifier community composition varied over sampling time. Although many clones were distantly related to the known NirS or NirK, some clones were related to the NirS from Burkholderiales and Rhodocyclales bacteria, and some were related to the NirK from Rhizobiales bacteria. These denitrifiers may play an important role in denitrification in the rice paddy field soil. The quantitative PCR results showed that nirK was more abundant than nirS in all soil samples, but the nirK/nirS ratio decreased after water logging. These results suggest that both diversity and quantity changed over time in the rice paddy field soil, in response to the soil condition.     

Relationships of culm surface area to other culm dimensions for bamboo, Phyllostachys pubescens

We determine the relationships of culm surface area to other culm dimensions for one of the largest bamboo species, Phyllostachys pubescens Mazel ex Houzeau de Lehaie. A total number of 150 sample culms were collected from a stand of P. pubescens in Mt. Toshima, Kumamoto Prefecture, western Japan. The culm surface area for each sample was calculated, and then the relationships of the culm surface area to basal area and product of diameter at breast height (dbh) and culm height were analyzed. The relationship between culm surface area and basal area could be described successfully by the power equation, whereas there was a linear relationship of culm surface area to product of dbh and height. Although the regression equations determined here would be useful in estimating culm surface area of P. pubescens, it is necessary to select an appropriate equation depending on the purpose and available time and labor.     

Antimicrobial activity against <Emphasis Type="Italic">Streptococcus sobrinus</Emphasis> and glucosyltransferase inhibitory activity of taxifolin and some flavanonol rhamnosides from kempas (<Emphasis Type="Italic">Koompassia malaccensis</Emphasis>) extracts

Twenty plant materials collected from the islands of Java and Kalimantan in Indonesia were extracted with 50% aqueous ethanol (crude extract). The crude extracts were assayed for antimicrobial activities against Streptococcus sobrinus and for glucosyltransferase (GTase) inhibition. Fourteen extracts inhibited the growth of S. sobrinus by more than 50% and six extracts inhibited GTase activity by more than 50% at a concentration of 100 μg/ml. Koompassia malaccensis (kempas) extracts showed 90% depression of S. sobrinus growth and 80% inhibition of GTase activity at a concentration of 100 μg/ml. Kempas crude extracts were subjected to column chromatography using Sephadex LH-20 and then preparative high-performance liquid chromatography to isolate four compounds A, B, C, and D. These compounds were identified as taxifolin and the flavanonol rhamnoside isomers neoastilbin, astilbin, and isoastilbin, respectively, from ¹H and ¹³C nuclear magnetic resonance (NMR) spectra and other two-dimensional NMR techniques (COSY, HMBC, and HMQC). Each compound depressed the growth of S. sobrinus over a concentration range of 9.3242.7 μg/ml and showed GTase inhibitory activity with IC₅₀ values in the range 27.4–57.3 μg/ml. Taxifolin and flavanonol rhamnoside isomers isolated for the first time from kempas could be potent compounds for preventing dental caries. Part of this report was presented at the 57th Annual Meeting of the Japan Wood Research Society Conference, Hiroshima, 2007     

First application of PCR-Luminex system for molecular diagnosis of fungicide resistance and species identification of fungal pathogens

Fungicide resistance in plant pathogens is often caused by a single point mutation in a gene encoding fungicide target proteins. Such is the case for resistance to MBI-D (inhibitors of scytalone dehydratase in melanin biosynthesis) fungicides in rice blast fungus (Magnaporthe oryzae), which is caused by a mutation in the scytalone dehydratase gene that results in a replacement of valine with methionine at codon 75 of the fungicide target protein. PCR-Luminex, a novel system developed for high-throughput analysis of single nucleotide polymorphisms (SNPs) was successfully introduced to diagnose MBI-D resistance using specific oligonucleotide probes coupled with fluorescent beads. The PCR-Luminex system was further tested for its potential in identifying species causing Fusarium head blight on wheat. Four major pathogens, Fusarium graminearum (=F. asiaticum), F. culmorum, F. avenaceum, and Microdochium nivale, known to cause the disease, were tested, and the species were identified using the PCR-Luminex method. So far, this report is the first on the application of the DNA-based PCR-Luminex system in the area of crop protection and/or agricultural sciences.     

Genetic analysis and PCR-based identification of major <Emphasis Type="Italic">Fusarium</Emphasis> species causing head blight on wheat in Japan

Identifying the Fusarium species cause Fusarium head blight (FHB) and produces mycotoxins in wheat and other cereal is difficult and time consuming because of confusing phenotypic classification systems. In Japan, the F. graminearum complex, F. culmorum, F. avenaceum, and Microdochium nivale predominantly cause FHB. The internal transcribed spacer (ITS) and 5.8S of rDNA, a partial sequence of β-tubulin and mitochondrial cytochrome b (cytb) genes of the four species were PCR-amplified and analyzed. On the basis of the ITS, β-tubulin and cytb sequences, F. avenaceum and M. nivale are distinct from the F. graminearum complex and F. culmorum, whereas the F. graminearum complex is closely related to F. culmorum. Moreover, thiophanate–methyl-resistant isolates of the F. graminearum complex and F. culmorum did not have an amino acid substitution at amino acid codon 198 or 200 of β-tubulin. In contrast, very highly or highly thiophanate–methyl-resistant isolates of M. nivale had Glu (GAG) substituted with Ala (GCG) or Lys (AAG) at codon 198, respectively. The allele-specific PCR assay was used to identify the F. graminearum complex and F. culmorum, and these Fusarium species could be distinguished rapidly.     

Ultrastructural study on acibenzolar-S-methyl-induced scab resistance in epidermal pectin layers of Japanese pear leaves

The infection behavior of Japanese pear scab pathogen Venturia nashicola race 1 was studied ultrastructurally in acibenzolar-S-methyl (ASM)-pretreated susceptible Japanese pear (cv. Kousui) leaves to determine the mechanism of ASM-induced scab resistance. On ASM-pretreated leaf surfaces, the infection behavior (conidial germination and appressorial formation) was similar to that on distilled water (DW)-pretreated leaves prior to cuticle penetration by the pathogen. However, after penetration, differentiated behavior was found in epidermal pectin layers and middle lamellae of the ASM-pretreated leaves. Subcuticular hyphae in epidermal pectin layers and middle lamellae of ASM-pretreated pear leaves were observed at lower frequency than in DW-treated leaves. The results indicated that fungal growth was suppressed in ASM-pretreated pear leaves. In the pectin layers of ASM- and DW-pretreated leaves, some hyphae showed morphological modifications, which were used as criteria to judge collapse of hyphal cells, including plasmolysis, necrotic cytoplasm, and cell wall destruction. More hyphae had collapsed in ASM-pretreated leaves than in DW-treated ones. In addition, the cell walls of collapsed hyphae broke into numerous fibrous and amorphous pieces, suggesting that ASM-induced scab resistance might be associated with cell-wall-degrading enzymes from pear plants. In addition, results from morphometrical analysis suggested that the activity or production of pectin-degrading enzyme from hyphae were inhibited by ASM application when compared with DW treatment.     

Effects of cooling rate on post-thaw motility and fertility of Japanese pearl oyster <Emphasis Type="Italic">Pinctada fucata martensii</Emphasis> spermatozoa

The post-thaw motility and fertility of Japanese pearl oyster sperm show large variances, even among sperm samples obtained from the same individuals. This study aimed to clarify the factors that cause such differences. Spermatozoa were diluted 50 times with diluent comprising 10 % methanol, 18 % fetal bovine serum, and 72 % seawater, and dispensed into 0.25 ml straws. A total of 59 straws were cooled, one by one, at 11 different heights from the surface of liquid nitrogen (LN) to −50 °C, and then immediately immersed in LN. After thawing the straws, the relationships between the cooling rate and the post-thaw motility and post-thaw fertility of the spermatozoa were examined. Both the post-thaw motility and the post-thaw fertility showed a sharp peak when the straws were cooled at around −20 °C/min. There was a strong correlation between post-thaw motility and fertility (P < 0.001). There was a large difference in the cooling rates and the post-thaw motilities and fertilities of the spermatozoa, even between straws cooled at the same height. These results indicate that the optimum range for the cooling rate of oyster spermatozoa is quite narrow, and the method of cooling straws at a fixed distance from the LN surface is unsuitable for the cryopreservation of Japanese pearl oyster spermatozoa.