Identification of defect position in a wooden beam from the power spectrum of longitudinal vibration

The inverse solution procedure that enables the identification of the defect position in a beam from the resonance frequency was exploited. Resonance frequency shifts of a power spectrum due to defects in a longitudinally vibrating beam when both ends are free were investigated by both numerical and experimental analysis. Calculation by a transfer matrix method showed that the frequency shift was large when the defect position coincides with a node of vibration and that no shift occurs when it coincides with a loop of vibration. The frequency shift could be approximated by a sinusoidal curve. Calculation results agreed well with those of the experiment in which artificial round holes were drilled as the defect model. Experimental equations predicting the amount of the frequency shift in function of the defect position were obtained. In the inverse procedure, the defect position was determined by comparing the resonance frequencies between the experimental and estimated power spectra so that the coincidence factor S(x) became a minimum. The results showed the validity of the proposed method to identify the defect positions of fewer than two predominant defects.     

Improved method for in situ biolistic transformation to analyze barley–powdery mildew interactions

Although the use of stable transformants is indispensable to elucidate mechanisms underlying molecular plant–pathogen interactions, this approach remains difficult to apply to crops. Alternatively, biolistic transformation has often been used as a transient expression method in various plants. In this study, we developed a method for in situ biolistic transformation without separating leaves from barley seedlings by using a hand-held particle bombardment system because unwounded leaves are preferable for analyzing interactions between barley and Blumeria graminis f. sp. hordei which requires healthy living cells. As a result, we found that the infection rate in intact leaves was higher than in separated leaves and that the transformation efficiencies in leaves were higher when plants were grown in vermiculite rather than in culture soil. Furthermore, we determined the appropriate inoculation time after bombardment to analyze the incompatible interaction and successfully monitored the gradual occurrence of cell death over time. Our system was suitable for relatively long-term follow-up analysis of the fate of each single cell during plant–pathogen interactions.     

Molecular characterization of the Fusarium graminearum species complex in Japan

Members of the Fusarium graminearum species complex are important cereal pathogens worldwide and belong to one of at least nine phylogenetically distinct species. We examined 298 strains of the F. graminearum species complex collected from wheat or barley in Japan to determine the species and trichothecene chemotype. Phylogenetic analyses and species-diagnostic polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLPs) revealed the presence and differential distribution of F. graminearum sensu stricto (s. str.) and F. asiaticum in Japan. F. graminearum s. str. is predominant in the north, especially in the Hokkaido area, while F. asiaticum is predominant in southern regions. In the Tohoku area, these species co-occurred. Trichothecene chemotyping of all strains by multiplex PCR revealed significantly different chemotype compositions of these species. All 50 strains of F. graminearum s. str. were of a 15- or 3-acetyl deoxynivalenol type, while 173 (70%) out of 246 strains of F. asiaticum were of a nivalenol type. The possibility of gene flow between the two species was investigated by use of 15 PCR-RFLP markers developed in this study. However, no obvious hybrids were detected from 98 strains examined, including strains collected from regions where both species co-occur.     

Monitoring by real-time PCR of three water-borne zoosporic Pythium species in potted flower and tomato greenhouses under hydroponic culture systems

The high-temperature-tolerant Pythium species P. aphanidermatum, P. helicoides, and P. myriotylum cause serious diseases in many crops under hydroponic culture systems in Japan. Control of the diseases is difficult because these zoosporic pathogens spread quickly. In this study, a real-time PCR method was developed for monitoring the spread of zoospores of the three pathogens. Specific primers and TaqMan probes were established using the internal transcribed spacer regions of the rDNA. Specificity was confirmed using known isolates of each species and closely related non-target species. The sensitivity of DNA detection was 10 f. for each pathogen. 10 f. DNA corresponded to 4 P. aphanidermatum, 3 P. myriotylum, and 4 P. helicoides zoospores, respectively. Therefore, this real-time PCR method was used to evaluate and monitor zoospores in the nutrient solutions of ebb-and-flow irrigation systems for potted flower production and closed hydroponic culture systems for tomato production. The results indicated that the pathogens were present in the hydroponic culture systems throughout the year, and spread before disease occurrence.     

Seedling blight of Glycyrrhiza uralensis caused by Pythium myriotylum,P. aphanidermatum and P. spinosum and identifying primary inoculum sources using multiplex PCR detection

Pythium species, isolated from seedlings of Glycyrrhiza uralensis with blight, were identified as P. myriotylum, P. aphanidermatum, and P. spinosum on the basis of morphological characteristics and sequences of the internal transcribed spacer regions of rDNA. In pathogenicity tests, the isolates of the three Pythium species caused blight, producing the original disease symptoms. The primary inoculum source was determined using a multiplex PCR to detect the pathogen. All the Pythium species were detected in the soils of fields with the diseased plants and in soils of adjacent field soils.     

Specific detection of Type 1 and Type 2 isolates of Pyrenochaeta lycopersici by loop-mediated isothermal amplification reaction

Pyrenochaeta lycopersici is the causal agent of corky root, an important soilborne disease of tomato (Lycopersicon esculentum) and other solanaceous crops. P. lycopersici isolates are classified into Types 1 and 2 on the basis of several physiological and molecular features. In this study, we aimed to establish a loop-mediated isothermal amplification (LAMP) method to identify Types 1 and 2 isolates. Using specific LAMP primer sets for Types 1 and 2, both types were easily identified within 35 min. The LAMP method demonstrated equal sensitivity to the polymerase chain reaction in molecular identification of the pathogen in cultured mycelia, infested plant roots, and their surrounding soil. The LAMP technology will contribute to the detection and identification of the pathogen, crop protection, and thorough understanding about the ecology of corky root disease of tomato.     

Morphological study in B16F10 murine melanoma cells after photodynamic hyperthermal therapy with indocyanine green (ICG)

Photodynamic hyperthermal therapy (PHT) with indocyanine green (ICG) is a combination of photodynamic therapy (PDT) and hyperthermia (HT). The low toxicity of ICG with an absorption wavelength of 700-800 nm is thought to make it a good candidate as a photosensitizer for PHT. Upon irradiation, ICG produces oxygen radicals and generates heat. The optimal concentration of ICG and the PHT post-irradiation time effects were evaluated by the cytotoxicity of the treatment on B16F10 murine melanoma. The cytotoxicity of PHT was determined based on the morphology of apoptotic and necrotic cells under phase-contrast microscope, confocal laser scanning microscope (CLSM) with DAPI and Annexin V-FITC/PI double staining, and cell surface structure evaluation by scanning electron microscopy (SEM). The use of ICG at a concentration of 150 μM was selected, as cell proliferation was inhibited from 0 to 24 hr post-PHT with a 3-fold decrease in cell viability (P<0.001) compared to the control group. A morphological observation revealed apoptotic and some degree of necrotic features in the PHT-treated cells.     

Inoculum effect of arbuscular mycorrhizal fungi on soybeans grown in long-term bare-fallowed field with low phosphate availability

ABSTRACT

Inoculation with arbuscular mycorrhizal fungi (AMF) can increase the growth of host plants, especially under condition of low phosphate (P) availability. Although this effect is shown relatively easily in simplified systems such as pot experiments, it is often hard to show in the field because of complicating factors such as competition with indigenous AMF. We conducted an AMF inoculation experiment with three Japanese soybean cultivars (Enrei, Misuzudaizu, and Akishirome) in an allophanic (Umbric Silandic) Andosol field under the long-term selective application of major nutrients (NPK and -P) and bare fallow. In the inoculation plots, introduced AMF were well colonized in soybean roots at flowering stage. In the -P plots, inoculation tended to increase the shoot dry weight of all the three soybean cultivars; this effect remained until harvest. Although a significant difference is not recognized, there was a tendency of residual effect on Enrei in the following year. In the NPK plots, inoculation did not significantly increase the shoot dry weight. We thought that in the -P plots, the long-term selective application of N and K and the long-term maintenance of bare fallow created the soil conditions of low P availability and poor native AMF. Thus, introduced AMF can benefit soybean growth and yield in the soil with low competitor AMF density and low phosphate availability.     

Characterization of a New Subgroup of Rhizoctonia solani Anastomosis Group 1 (AG-1-ID), Causal Agent of a Necrotic Leaf Spot on Coffee

ABSTRACT A new foliar disease on coffee leaves was observed in Mindanao, Philippines, in 1996. The symptoms appeared as large circular or irregularly shaped necrotic areas with small circular necrotic spots (1 mm or less in diameter) usually found around the periphery of the large necrotic areas. Rhizoctonia solani was consistently isolated from these diseased coffee leaves. Isolates obtained were multinucleate (3 to 12 nuclei per hyphal cell), had an optimum temperature for hyphal growth at 25 degrees C, prototrophic for thiamine, and anastomosed with tester isolates belonging to R. solani anastomosis group 1 (AG-1). Mature cultures on potato dextrose agar (PDA) were light to dark brown. Sclerotia, light brown to brown, were formed on the surface of PDA and covered the whole mature colony culture. Individual sclerotia often aggregated into large clumps (3 to 8 mm in diameter) and their color was brown to dark brown. In pathogenicity tests, isolates from coffee caused necrotic symptoms on coffee leaves, whereas isolates of AG-1-IA (not isolated from coffee), 1-IB, and 1-IC did not. The results of analyses of restriction fragment length polymorphism of ribosomal DNA internal transcribed spacer, random amplified polymorphism DNA, and fatty acid profiles showed that R. solani isolates from coffee are a population of AG-1 different from AG-1-IA, 1-IB, and 1-IC. These results suggest that R. solani isolates from coffee represent a new subgroup distinct from AG-1-IA, 1-IB, and 1-IC. A new subgroup ID (AG-1-ID) is proposed.     

Changes in heart rate variability in horses during immersion in warm springwater

OBJECTIVE: To determine the effects of immersion in warm springwater (38 degrees to 40 degrees C) on autonomic nervous activity in horses. ANIMALS: 10 male Thoroughbreds. PROCEDURE: Electrocardiograms were recorded from horses for 15 minutes during a warm springwater bath after being recorded for 15 minutes during stall rest. Variations in heart rate (HR) were evaluated from the power spectrum in terms of low frequency (LF, 0.01 to 0.07 Hz) power and high frequency (HF, 0.07 to 0.6 Hz) power as indices of autonomic nervous activity. RESULTS: Mean (+/- SE) HR during stall rest and immersion in warm springwater was 31.1 +/- 1.7 and 30.3 +/- 1.0 beat/min, respectively. No significant difference was found between the HR recorded during stall rest and that recorded during immersion in warm springwater. The HF power significantly increased from 1,361 +/- 466 milliseconds2 during stall rest to 2,344 +/- 720 milliseconds2 during immersion in warm springwater. The LF power during stall rest and immersion in warm springwater was 3,847 +/- 663 and 5,120 +/- 1,094 milliseconds2, respectively, and were not significantly different from each other. Similarly, the LF:HF ratio did not change during immersion in warm springwater. The frequency of second-degree atrioventricular block, which was observed in 2 horses, increased during immersion in warm springwater, compared with during stall rest. CONCLUSIONS AND CLINICAL RELEVANCE: Increases in HF power indicates that the parasympathetic nervous activity in horses increases during immersion in warm springwater. Thus, immersion in warm springwater may provide a means of relaxation for horses.