Visualization of latent fingerprints using fluorescence lifetime imaging on paper emitting strong fluorescence.

Latent fingerprints were successfully visualized using fluorescence lifetime imaging (FLIM) on paper which emits strong fluorescence with a lifetime close to that of fingerprints and thus from which it is difficult for time-resolved spectroscopy to visualize fingerprints. Latent fingerprint samples on paper were excited using a 450 nm or 532 nm nanosecond pulsed-laser, and time-resolved fluorescence images were obtained at a delay time of 6-16 ns in intervals of 1 ns, to the excitation pulse. The excitation beam was expanded using a lens, and the fluorescence from the fingerprints was captured using an intensified CCD camera. Because of the large fluorescence intensity of the background paper of approximately two to four orders of magnitude larger than that of the fingerprint, the fingerprint was not visualized on each fluorescence image by time-resolved spectroscopy. However, the fingerprint was visualized in a FLIM image constructed using a series of the fluorescence images for the case with the fluorescence intensity of the background paper being four orders of magnitude larger than that of the fingerprint. The difference in fluorescence lifetime in the FLIM image of the visualized fingerprint and background paper was in the order of 0.1 ns, which was an order of magnitude smaller than the inherent fluorescence lifetime of a few nanoseconds for the fingerprints and paper. It was demonstrated that, at a background fluorescence intensity with a certain order of magnitude larger than that of fingerprints, FLIM has the potential to visualize latent fingerprints which cannot be visualized by time-resolved spectroscopy.

d-Tartrate utilization correlates with phylogenetic subclade in Pseudomonas cichorii.

Pseudomonas cichorii is divided into two subclades based on the 16S ribosomal RNA gene sequence and core genome multilocus sequence typing. It was shown that subclade 2 strains utilize d-tartrate as a sole carbon source, whereas subclade 1 strains do not. Draft genome sequencing was performed with P. cichorii strains to identify d-tartrate utilization genes. By genome comparative and homology search studies, an ∼7.1-kb region was identified to be involved in d-tartrate utilization. The region is subclade 2 specific, and contains tarD and dctA genes, which encode a putative enzyme and transporter of d-tartrate, respectively. When the region was introduced into subclade 1 strains, the transformants were able to utilize d-tartrate. Partial fragments of tarD and dctA were amplified from all subclade 2 strains tested in this study by PCR using gene-specific primers, but not from subclade 1 strains. This is the first report on the genetic analysis of biochemical characteristics corresponding to a specific phylogenetic group in P. cichorii.

Complete Genome Sequences of Ralstonia solanacearum Strains Isolated from Zingiberaceae Plants in Japan.

Here, we report the complete genome sequences of three Ralstonia solanacearum strains isolated from Zingiberaceae plants in Japan. The total genome sizes of these strains ranged from 5.87 to 6.05 Mb. Strains MAFF 211472, MAFF 211479, and MAFF 311693 each carried one chromosome and one megaplasmid. MAFF 311693 contained an additional 71.9-kb plasmid.

Visualization of Aged Fingerprints with an Ultraviolet Laser.

Detection of aged fingerprints is difficult because they can degrade over time with exposure to light, moisture, and temperature. In this study, aging fingerprints were visualized by time-resolved spectroscopy with an ultraviolet-pulsed laser. Fingerprints were prepared on glass slides and paper and then stored under three lighting conditions and two humidity conditions for up to a year. The fluorescence intensities of the fingerprints decreased with time. Samples were stored in the dark degraded less than in sunlight or under a fluorescent lamp. Samples were stored under low humidity degraded less than under moderate humidity. As the storage period increased, a fluorescence emission peak appeared that was at a longer wavelength than the peak visible in earlier spectra. This peak was used for visualization of an aged fingerprint over time. An image of the fingerprint was not initially visible, but an image appeared as the time since deposition of the fingerprint increased.

Species-Specific Detection of Mycosphaerella polygoni-cuspidati as a Biological Control Agent for Fallopia japonica by PCR Assay.

The ascomycete fungus Mycosphaerella polygoni-cuspidati has been undergoing evaluation as a potential classical biological control agent for the invasive weed Fallopia japonica (Japanese knotweed), which has become troublesome in Europe and North America. In advance of the potential release of a biocontrol agent into a new environment, it is crucial to develop an effective monitoring system to enable the evaluation of agent establishment and dispersal within the target host population, as well as any potential attacks on non-target species. Therefore, a primer pair was designed for direct, rapid, and specific detection of the Japanese knotweed pathogen M. polygoni-cuspidati based on the sequences of the internal transcribed spacer regions including the 5.8S rDNA. A PCR product of approximately 298 bp was obtained only when the DNA extracted from mycelial fragments of M. polygoni-cuspidati was used. The lower limit of detection of the PCR method was 100 fg of genomic DNA. Using the specific primer pair, M. polygoni-cuspidati could be detected from both naturally and artificially infected Japanese knotweed plants. No amplification was observed for other Mycosphaerella spp. or fungal endophytes isolated from F. japonica. The designed primer pair is thus effective for the specific detection of M. polygoni-cuspidati in planta.

A single amino acid at N-terminal region of the 2b protein of cucumber mosaic virus strain m1 has a pivotal role in virus attenuation.

Host responses to infection by a mild strain of cucumber mosaic virus, termed CMV-m1, were re-examined in several plant species in comparison with those by a severe strain CMV-Y. Mild systemic symptoms were developed on the six plant species inoculated with CMV-m1. Virus titer in the Nicotiana benthamiana plants infected with CMV-m1 was significantly lower than those infected with CMV-Y, although infection by CMV-m1 interfered with further infection by CMV-Y in the plants. Subsequently, the attenuated virulence of CMV-m1 was analyzed by reassortment and recombination analyses between CMV-m1 and CMV-Y RNAs. The results suggested that the 2b protein of CMV-m1 (m1-2b) is involved in the formation of mild symptoms in N. benthamiana. Furthermore, site-directed mutagenesis demonstrated that Thr18 of m1-2b is responsible for formation of mild symptoms. Local RNA silencing suppressor activity of m1-2b was a little lower than that of severe strain CMV-Y. We discuss the relationship between attenuation of CMV-m1 and the features of m1-2b.

Induction of antiviral responses by acibenzolar-s-methyl against cucurbit chlorotic yellows virus in Melon.

Cucurbit chlorotic yellows virus (CCYV) (family Closteroviridae, genus Crinivirus) is an emerging virus which causes severe diseases on melon (Cucumis melo) plants. CCYV-infected melon plants display yellowing, mottling, chlorosis, or chlorotic spots on leaves. To develop a new control strategy, the potential for 1,2,3-benzothiadiazole-7-thiocarboxylic acid-S-methyl-ester (ASM) to suppress CCYV infection was evaluated. ASM treatment on melon plants greatly increased the expression levels of pathogenesis-related 1a gene, a marker gene for systemic acquired resistance. ASM treatment on melon plants before inoculation of CCYV suppressed systemic symptoms and decreased CCYV accumulation. ASM treatment on melon even after inoculation of CCYV reduced disease severity and accumulation levels of CCYV. The results show the potential for ASM treatment on attenuation of the CCYV disease symptoms.

Comparison of liquid crystal display monitors calibrated with gray-scale standard display function and with γ 2.2 and iPad: observer performance in detection of cerebral infarction on brain CT.

OBJECTIVE: The purpose of the study was to compare observer performance in the detection of cerebral infarction on a brain CT using medical-grade liquid crystal display (LCD) monitors calibrated with the gray-scale standard display function and with γ 2.2 and using an iPad with a simulated screen setting. MATERIALS AND METHODS: We amassed 97 sample sets, from 47 patients with proven cerebral infarction and 50 healthy control subjects. Nine radiologists independently assessed brain CT on a gray-scale standard display function LCD, a γ 2.2 LCD, and an iPad in random order over 4-week intervals. Receiver operating characteristic (ROC) analysis was performed by using the continuous scale, and the area under the ROC curve (A(z)) was calculated for each monitor. RESULTS: The A(z) values for gray-scale standard display function LCD, γ 2.2 LCD, and iPad were 0.875, 0.884, and 0.839, respectively. The difference among the three monitors was very small. There was no significant difference between gray-scale standard display function LCD and γ 2.2 LCD. However, the A(z) value was statistically significantly smaller for the iPad than the γ 2.2 LCD (p < 0.05). CONCLUSION: Observer performance for detecting cerebral infarction on the LCD with γ 2.2 calibration was found to be similar to the LCD with gray-scale standard display function calibration. Although observer performance using the iPad was poorer than that using the other LCDs, the difference was small. Therefore, the iPad could not substitute for other LCD monitors. However, owing to the promising potential advantages of tablet PCs, such as portability, further examination is needed into the clinical use of tablet PCs.

What causes false-negative PET findings for solid-type lung cancer?

BACKGROUND: To investigate possible causes for false-negative findings on PET scans for solid-type lung cancers, we retrospectively compared PET findings to clinical and pathological features using multivariate analysis. METHODS: We reviewed PET/CT records, clinical records, preoperative thin-section CT images, and postoperative pathological records and selected only solid-type primary lung cancers with lesions ≤ 40 mm in diameter that had been definitively diagnosed by surgical resection. PET images with SUVmax of ≥ 2.5 were considered PET-positive. Logistic regression analysis was used to identify independent predictors of PET-positive or negative among five factors: body weight, blood glucose level, lesion size, location, and histological classification. RESULTS: A total of 187 solid-type primary lung cancers were selected. Forty lesions (21.4%) were judged as PET-negative and 147 lesions (78.6%) were judged as PET-positive. Multivariate logistic analysis for the 187 lesions revealed that lesion size (p<0.001) and histological tumour type (p<0.001) were significant factors for determining whether PET findings were negative. CONCLUSIONS: Among solid-type lung cancers, lesion size and histopathological findings were significantly associated with FDG uptake. In particular, it warrants attention that lesions ≤ 2 cm and bronchioloalveolar carcinoma and well-differentiated adenocarcinoma have a tendency for negative PET findings.

Endophytic fungi associated with Fallopia japonica (Polygonaceae) in Japan and their interactions with Puccinia polygoni-amphibii var. tovariae, a candidate for classical biological control.

Fallopia japonica (Polygonaceae), or Japanese knotweed, is now spreading globally, causing serious problems in Europe and North America in both natural and urban habitats. There is an urgent need for alternative management solutions, and classical biological control, using coevolved natural enemies found in the native range, is currently being investigated. Here, we isolated fungal endophytes from F. japonica in Japan, its natural habitat, to find endophytes that might increase the virulence of a coevolved rust pathogen, Puccinia polygoni-amphibii var. tovariae. A total of 1581 fungal endophytes were recovered from F. japonica and classified into 15 taxa. Five genera (Colletotrichum, Pestalotiopsis, Phoma, Phomopsis, and Alternaria) were dominant as endophytes in F. japonica. A greenhouse study of the dominant endophyte-pathogen interactions revealed three types of reactions: suppressive, synergistic, and neutral. In particular, one Phomopsis isolate--closely related to Diaporthe medusaea, based on ITS sequences--promoted the pathogenic aggressiveness of P. polygoni-amphibii var. tovariae and, therefore, this interaction is potentially useful to increase the effectiveness of the rust fungus as a biological control agent of F. japonica in its invasive range.

Genetic diversity and ecological evaluation of fluorescent pseudomonads isolated from the leaves and roots of potato plants.

A total of 828 isolates of fluorescent pseudomonads (FPs) were obtained from the leaves (305 isolates) and roots (523 isolates) of potato plants grown in different geographical locations in Japan, and 16S rRNA gene sequences of 776 isolates were successfully determined by direct PCR sequencing. Clustering analysis (≥99% identity) identified 13 and 26 operational taxonomic units (OTUs) for leaf- and root-associated FPs, respectively, and 29 OTUs were identified in the phytosphere of potato plants. Among them, 7 and 9 OTUs showed a significantly biased distribution to the leaves and roots, respectively. Phylogenetic analysis revealed that 3 dominant OTUs for leaf-associated FPs were grouped in a cluster of leaf-associated pathogens, such as Pseudomonas cichorii and Pseudomonas viridiflava. In contrast, 4 OTUs were located in a cluster of saprophytic pseudomonads. Among them, 3 OTUs showed high similarity to Pseudomonas koreensis and Pseudomonas vancouverensis, both of which have been reported to be beneficial for biological control or plant growth promotion. These data provide key information for efficient surveying and utilization of beneficial FPs in agricultural practices.

FDG-PET performed concurrently with initial I-131 ablation for differentiated thyroid cancer.

OBJECTIVE: Differentiated thyroid cancers (DTCs) are commonly treated by total thyroidectomy followed by I-131 radioiodine ablation to eradicate any residual thyroid tissue and to detect any metastatic lesions on post-treatment whole body scans (TxWBS). However, some DTCs do not trap iodine, resulting in negative whole body scanning. Fluorine-18-fluorodeoxyglucose positron emission tomography (FDG-PET) has proven to be a valuable diagnostic technique for detecting many types of malignant tumors and metastases. The purpose of this study was to evaluate FDG-PET performed concurrently with initial I-131 ablation for its ability to detect lymph node metastasis and for its role in the management of DTC patients. METHODS: A total of 54 patients (16 males and 38 females; median age = 50 years) with histologically proven DTC underwent both FDG-PET and subsequent I-131 ablation. A dose of 3.7 GBq I-131 was administered to 51 patients, 2.96 GBq was administered to 1 patient, and 2.22 GBq was administered to 2 patients. FDG-PET or PET/CT was performed 3-4 days prior to ablation. TxWBS was conducted 1 week after therapy. FDG-PET scans and TxWBS were interpreted by consensus of 2 experienced radiologists. Serum thyroglobulin (Tg) levels at 3-6 months after ablation were compared between PET-positive and PET-negative patients. RESULTS: FDG-PET was positive in 25 sites (thyroid bed: n = 9; cervical lymph nodes: n = 12; mediastinal lymph nodes: n = 3; and axillary lymph nodes: n = 1) of 18 patients (33%). Only 5 of 16 lymph nodes (31%) that were PET-positive were also positive on TxWBS. The success rate of Tg-negative after ablation was significantly lower for patients with PET-positive scans than for those with PET-negative scans (p = 0.026). CONCLUSIONS: FDG-PET performed concurrently with I-131 ablation can detect lymph node metastases in which radioiodine does not accumulate and may influence the management and treatment options for DTC patients.

Infection dynamics in viral spread and interference under the synergism between Cucumber mosaic virus and Turnip mosaic virus.

Mixed infection of Cucumber mosaic virus (CMV) and Turnip mosaic virus (TuMV) induced more severe symptoms on Nicotiana benthamiana than single infection. To dissect the relationships between spatial infection patterns and the 2b protein (2b) of CMV in single or mixed infections, the CMV vectors expressing enhanced green fluorescent or Discosoma sp. red fluorescent proteins (EGFP [EG] or DsRed2 [Ds], respectively were constructed from the same wild-type CMV-Y and used for inoculation onto N. benthamiana. CMV2-A1 vector (C2-A1 [A1]) has a functional 2b while CMV-H1 vector (C2-H1 [H1]) is 2b deficient. As we expected from the 2b function as an RNA silencing suppressor (RSS), in a single infection, A1Ds retained a high level of accumulation at initial infection sites and showed extensive fluorescence in upper, noninoculated leaves, whereas H1Ds disappeared rapidly at initial infection sites and could not spread efficiently in upper, noninoculated leaf tissues. In various mixed infections, we found two phenomena providing novel insights into the relationships among RSS, viral synergism, and interference. First, H1Ds could not spread efficiently from vasculature into nonvascular tissues with or without TuMV, suggesting that RNA silencing was not involved in CMV unloading from vasculature. These results indicated that 2b could promote CMV to unload from vasculature into nonvascular tissues, and that this 2b function might be independent of its RSS activity. Second, we detected spatial interference (local interference) between A1Ds and A1EG in mixed infection with TuMV, between A1Ds (or H1Ds) and TuMV, and between H1Ds and H1EG. This observation suggested that local interference between two viruses was established even in the synergism between CMV and TuMV and, again, RNA silencing did not seem to contribute greatly to this phenomenon.

Diversity of culturable chitinolytic bacteria from rhizospheres of agronomic plants in Japan.

A total of 100 isolates of chitinolytic bacteria were obtained from the rhizospheres of various agronomic plants, and the 16S rRNA gene sequences of these isolates were determined. Phylogenetic analyses revealed that 81 isolates belonged to the classes Betaproteobacteria (39 isolates) and Gammaproteobacteria (42 isolates). Of the remaining 19 isolates, 16 belonged to the phylum Firmicutes. Clustering analysis identified 6 and 3 operational taxonomic units (OTUs) in Gammaproteobacteria and Betaproteobacteria, respectively, at the genus level. The majority of chitinolytic bacteria in Gammaproteobacteria belonged to the genera Serratia, Stenotrophomonas, and Lysobacter (14, 15, and 7 isolates, respectively) while those in Betaproteobacteria belonged to the genus Mitsuaria (37 isolates). The 16 isolates placed in Firmicutes belonged to 2 genera, Paenibacillus and Bacillus (8 isolates each). The isolates in the remaining OTUs belonged to the genera Erwinia, Aeromonas, Pseudomonas, Achromobacter, Flavobacterium, and Microbacterium, in less abundance. These results showed a wide distribution of culturable chitinolytic bacteria in the rhizospheres of various agronomic plants. Considering the potential antagonistic activity of chitinolytic enzymes against phytopathogenic fungi, which is exhibited by fungal cell wall degradation, the above-mentioned native chitinolytic bacteria in rhizospheres could potentially be utilized for the biological control of soil-borne phytopathogenic fungi.

Method for simple and rapid enumeration of total epiphytic bacteria in the washing solution of rice plants.

The phyllosphere is one of the most common habitats for terrestrial bacteria. However, little is known about the populations of bacteria, including unculturable bacteria, that thrive on plant surfaces. Here, we developed a fluorescent nuclear staining technique to easily and rapidly observe and enumerate populations of total and living epiphytic bacteria, with particular emphasis on the concentration by centrifugation and fixation of the epiphytic bacteria. An investigation on the optimal conditions for centrifugation and fixation revealed that centrifugation at 20 400g for 2 min and fixation with 0.5% glutaraldehyde solution were the optimum conditions for observation of the bacteria. Using this technique, we assessed the populations of the total and living bacteria on the surface of rice plants. When epiphytic bacteria were recovered from rice seeds (Oryza sativa 'Koshihikari'), the number of total and living bacterial cells was 7.36 and 6.85 log10·g⁻¹ (fresh mass) in the seed washing, respectively. In contrast, the numbers of total and living bacterial cells in the leaf sheath washings were 5.5-5.8 and 5.3-5.7 log10·g⁻¹, respectively. Approximately 5%-30% of the total bacteria in the washing solution of rice plant were culturable. The usefulness of the enumeration method and the amount of bacteria on the plant surfaces are discussed.

Impact of a defective RNA 3 from cucumber mosaic virus on helper virus infection dynamics.

D RNA 3Yalpha (D3Yalpha), a defective (D) RNA 3 derived from the Y strain of cucumber mosaic virus (CMV-Y), was further characterized in combination with different helper viruses in the genus Cucumovirus. Interestingly, Nicotiana benthamiana plants inoculated with CMV-D8 and D3Yalpha developed systemic symptoms which were different from those induced by CMV-D8. To elucidate the potential effects of D RNA 3 on virus infection on the basis of the original combination of CMV-Y and D3Yalpha, a point mutation was made in the coat protein gene, which determined symptoms, of either CMV-Y RNA 3 (Y3) or D3Yalpha. Symptoms induced on N. benthamiana and N. tabacum plants, and semi-quantitative RT-PCR revealed that the ratio of RNA 3 to D RNA 3 was associated with the differences of symptoms in the leaf tissues. Furthermore, analysis of in situ hybridization suggested that there were spatial effects between coat proteins of Y3 and D3Yalpha in the infected leaves.

Performance of the micro-PIC gaseous area detector in small-angle X-ray scattering experiments.

The application of a two-dimensional photon-counting detector based on a micro-pixel gas chamber (micro-PIC) to high-resolution small-angle X-ray scattering (SAXS), and its performance, are reported. The micro-PIC is a micro-pattern gaseous detector fabricated by printed circuit board technology. This article describes the performance of the micro-PIC in SAXS experiments at SPring-8. A dynamic range of >10(5) was obtained for X-ray scattering from a polystyrene sphere solution. A maximum counting rate of up to 5 MHz was observed with good linearity and without saturation. For a diffraction pattern of collagen, weak peaks were observed in the high-angle region in one accumulation of photons.

Characterization of a defective RNA derived from RNA 3 of the Y strain of cucumber mosaic virus.

A defective (D) RNA 3 naturally generated from the Y strain of cucumber mosaic virus (CMV-Y) was characterized using a biologically active cDNA clone. Sequencing of the clone revealed that the D RNA 3, named D RNA 3Yalpha, was derived from CMV-Y RNA 3 and contained a 10 nt deletion in the 5' untranslated region and a 162 nt deletion within the 3a open reading frame. Co-inoculation of D RNA 3Yalpha with CMV-Y derived from in vitro transcripts facilitated propagation of CMV-Y containing D RNA 3Yalpha in Nicotiana benthamiana or Nicotiana tabacum plants. CMV-Y, however, did not produce deletion mutants upon serial mechanical passages in the plants.

Co-inoculation of an antibiotic-producing bacterium and a lytic enzyme-producing bacterium for the biocontrol of tomato wilt caused by Fusarium oxysporum f. sp. lycopersici.

The antifungal compound 2,4-diacetylphloroglucinol-producing bacterium, Pseudomonas fluorescens strain LRB3W1, inhibits the growth of Fusarium oxysporum f. sp. lycopersici, and controls Fusarium wilt of tomato caused by F. oxysporum f. sp. lycopersici. On the other hand, Serratia marcescens strain B2, which produces cell wall-degrading enzyme chitinases, did not inhibit fungal growth and the suppressive effect of strain B2 against tomato Fusarium wilt was less than that of strain LRB3W1. Combined inoculation of strain LRB3W1 with strain B2 was more effective than treatment with strain LRB3W1 alone. When 2,4-diacetylphloroglucinol and the chitinolytic enzymes were applied in combination, a synergistic inhibitory effect against the pathogen was observed. It was possible that bacteria which produce cell wall-degrading enzymes enhanced the biocontrol effect of the antibiotic-producing bacterium against tomato Fusarium wilt.