Inhibitory effect of black cumin (Nigella sativa) seed essential oil on Fusobacterium nucleatum L-methionine-γ-lyase (L-methioninase) activity.

The enzyme L-methionine-γ-lyase is commonly found in a wide range of bacteria and catalyzes the α-elimination and γ-elimination of L-methionine to produce methyl mercaptan, α-ketobutyrate and ammonia. Black cumin seed essential oil (BC oil) reportedly exhibits deodorizing activity against methyl mercaptan. Therefore, we hypothesized that BC oil may also suppress methyl mercaptan production. In this study, we aimed to evaluate the inhibitory effect of BC oil on L-methionine-γ-lyase activity in Fusobacterium nucleatum. Recombinant L-methionine-γ-lyase was incubated under appropriate conditions with BC oil and its constituent thymoquinone. To analyze L-methionine-γ-lyase activity, α-ketobutyric acid and ammonia concentrations were determined. The concentrations of α-ketobutyric acid and ammonia were significantly decreased by 10 µg mL-1 of BC oil (P < 0.01) and 16.4 µg/mL of thymoquinone (P < 0.05). An enzyme kinetic assay showed a mixed inhibition pattern between L-methionine-γ-lyase and thymoquinone. In conclusion, BC oil not only had a deodorizing effect against methyl mercaptan but also an inhibitory effect on methyl mercaptan production through the suppression of L-methionine-γ-lyase activity. Thymoquinone may be mainly responsible for these effects of BC oil. Thus, application of natural BC oil may be adapted not only for medical use but also in other areas of life.

Effect of thymoquinone on Fusobacterium nucleatum­associated biofilm and inflammation.

Periodontitis affects oral tissues and induces systemic inflammation, which increases the risk of cardiovascular disease and metabolic syndrome. Subgingival plaque accumulation is a trigger of periodontitis. Fusobacterium nucleatum (FN) contributes to subgingival biofilm complexity by intercalating with early and late bacterial colonizers on tooth surfaces. In addition, inflammatory responses to FN are associated with the progression of periodontitis. Nigella sativa Lin. seed, which is known as black cumin (BC), has been used as a herbal medicine to treat ailments such as asthma and infectious diseases. The current study examined the inhibitory effect of BC oil and its active constituents, thymol (TM) and thymoquinone (TQ), on FN­associated biofilm and inflammation. FN­containing biofilms were prepared by co­cultivation with an early dental colonizer, Actinomyces naeslundii (AN). The stability and biomass of FN/AN dual species biofilms were significantly higher compared with FN alone. This effect was retained even with prefixed cells, indicating that FN/AN co­aggregation is mediated by physicochemical interactions with cell surface molecules. FN/AN biofilm formation was significantly inhibited by 0.1% TM or TQ. Confocal laser scanning microscopy indicated that treatment of preformed FN/AN biofilm with 0.01% of BC, TM or TQ significantly reduced biofilm thickness, and TQ demonstrated a cleansing effect equivalent to that of isopropyl methylphenol. TQ dose­dependently suppressed TNF­α production from a human monocytic cell line, THP­1 exposed to FN, yet showed no toxicity to THP­1 cells. These results indicated that oral hygiene care using TQ could reduce FN­associated biofilm and inflammation in gingival tissue.

Purification of a High Molecular Mass Protein in Streptococcus mutans.

Elucidation of a gene's function typically involves comparison of phenotypic traits of wild-type strains and strains in which the gene of interest has been disrupted. Loss of function following gene disruption is subsequently restored by exogenous addition of the product of the disrupted gene. This helps to determine the function of the gene. A method previously described involves generating a gtfC gene-disrupted Streptococcus mutans strain. Here, an undemanding method is described for purifying the gtfC gene product from the newly generated S. mutans strain following the gene disruption. It involves the addition of a polyhistidine-coding sequence at the 3' end of the gene of interest, which allows simple purification of the gene product using immobilized metal affinity chromatography. No enzymatic reactions other than PCR are required for the genetic modification in this method. The restoration of the gene product by exogenous addition after gene disruption is an efficient method for determining gene function, which may also be adapted to different species.

Method for functional analysis of a gene of interest in Streptococcus mutans: gene disruption followed by purification of a polyhistidine-tagged gene product.

Typical methods for elucidating the function of a particular gene involve comparative phenotypic analysis of the wild-type strain and a strain in which the gene of interest has been disrupted. We previously described a simple method for the generation of a gene-disrupted strain in Streptococcus mutans by replacing the gene of interest with an antibiotic resistance marker gene. It is also crucial that the function lost following the gene disruption is restored by exogenous addition of the gene product, but purification of this product can be difficult and involve a complex series of steps. In this study, we describe a simple method for the purification of gene products following gene disruption in S. mutans. The method involves the expression of an additional polyhistidine tag at the C-terminus of the gene product. The target protein can be simply purified by immobilized metal affinity chromatography and applied to a restoration assay. This method utilizes the genomes of both the wild-type strain and the gene-disrupted strain as PCR templates to generate the DNA construct. Therefore, generation of the gene-disrupted strain is a prerequisite for the present procedure. The combination of gene disruption and gene product purification results in an efficient method for the analysis of gene function that could be further adapted to various other bacterial species.

Simulations tackle abrupt massive migrations of energetic beam ions in a tokamak plasma.

In the late 1990s, fusion scientists at the Japanese tokamak JT-60U discovered abrupt large-amplitude events during beam-driven deuterium plasma experiments. A large spike in the magnetic fluctuation signal followed by a drop in the neutron emission rate indicates that energetic ions abruptly migrate out of the plasma core during an intense burst of Alfvén waves that lasts only 0.3 ms. With continued beam injection, the energetic ion population recovers until the next event occurs 40-60 ms later. Here we present results from simulations that successfully reproduce multiple migration cycles and report numerical and experimental evidence for the multi-mode nature of these intermittent phenomena. Moreover, we elucidate the role of collisional slow-down and show that the large-amplitude Alfvénic fluctuations can drive magnetic reconnection and induce macroscopic magnetic islands. In this way, our simulations allow us to gradually unravel the underlying physical processes and develop predictive capabilities.

Enhancement of oral moisture using tablets containing extract of Capparis masaikai Levl.

AIM OF THE STUDY: This study was performed to examine the effects of tablets containing an extract of Capparis masaikai Levl. (M-tablets) on enhancing oral moisture. SUBJECTS AND METHODS: The moistening effect of M-tablets was examined in 21 healthy subjects aged 25.1+/-2.4 (mean+/-S.D.) years in comparison with control tablets. After sucking tablets, the oral moisture was measured using a saliva wetness tester and a moisture checker. To evaluate the effects of the M-tablets on oral conditions, additional 50 subjects aged 30.6+/-7.5 years were examined. The subjects recorded changes in refreshment, oral moisture, ease in speaking, and taste of water using a visual analog scale (VAS). RESULTS: The L-SALIVO value of the M-tablet increased significantly from 1.83+/-0.17 (mean+/-S.E.M.) at baseline to 3.02+/-0.21 at 15 min (P<0.01). The Mucus((R)) value of the M-tablet also increased from 37.50+/-0.22 at baseline to 38.30+/-0.26 at 15 min (P<0.01). The VAS value for oral moisture increased significantly from 47.4+/-2.0 to 69.6+/-2.2 after taking the M-tablet (P<0.01). The VAS value for taste of water also increased from 50.0+/-1.1 to 66.7+/-3.2 (P<0.01). CONCLUSION: These results suggest that M-tablets are useful for enhancing oral moisture, which leads to improvement of oral conditions.

Absolute calibration of microfission chamber in JT-60U.

We have conducted calibrations of a microfission chamber, which was installed between the vacuum vessel and the toroidal field coils, by both a Cf-252 neutron source and real deuterium plasmas in JT-60U. The detector employs both pulse counting and Campbell (mean square voltage) modes in the electronics to cover a wide dynamic range of the neutron source strength. The pulse counting and Campbell modes were calibrated by Cf-252 and deuterium plasmas, respectively. Point efficiencies, counts per neutron from a point at a single angle, were measured for 27 locations of the neutron point source in toroidal scan. The efficiencies were influenced by the various components such as the vacuum vessel, port, and graphite tiles. The point efficiencies can be integrated and averaged with angle to provide toroidal line efficiencies. The line efficiencies of the microfission chamber and the nearest neutron monitor of the U-235 fission chamber were 5.38x10(-9) and 1.77x10(-8), respectively. Then the calibration for the Campbell mode was also performed by using a real deuterium plasma. The detection efficiency in the Campbell mode was about three-tenths of that of the nearest neutron monitor, which is consistent with the calibration result obtained by using a Cf-252 neutron source for pulse counting mode.

Design of microfission chamber for ITER operations.

Microfission chambers (MFCs) are one of the most important diagnostics for measuring neutron source strength in ITER. Using MFCs for high-power operations (fusion power of 100 kW-1 GW) and for low-power operations (<100 kW) in combination is one way to fulfill the target measurement requirements of ITER. The MFCs for high-power operations will be installed behind blanket modules in both the upper and lower outboard regions of the vacuum vessel so as to be insensitive to changes in the position of the plasma. For low-power operations, one possible location of MFCs is inside the equatorial (EQ) port. The effect of streaming neutrons and of changes in the position of the plasma on the responses of MFCs is estimated based on a neutron Monte Carlo calculation using the MCNP Version 5 code. Results suggest that the effect of streaming neutrons should be taken into account if the MFCs for high-power operations are installed closer than 20 cm to the gap between blanket modules. It has also been found for MFCs of low-power operations that the averaged output of the MFCs installed at the top and bottom of the EQ port is sensitive to horizontal plasma shifts but not to vertical shifts. This finding suggests that corrections based on the position of the plasma center will be needed for the absolute measurement of neutron source strength.

Usefulness of new wetness tester for diagnosis of dry mouth in disabled patients.

OBJECTIVES: The condition of dry mouth is an influential factor in the incidence of caries, periodontal disease, fungal infections, masticatory dysfunctions and denture function. Bedridden elderly and disabled persons often suffer from oral dryness and the aim of this study was to evaluate the usefulness of measuring the amount of moisture in the oral mucosa for clinical diagnosis of dry mouth in this group. SUBJECTS AND METHODS: The subjects were 20 elderly bedridden individuals, age range 65-89 years old, living in a nursing home and six healthy laboratory researchers, aged 20-46 years old, used as controls. Tongue dorsum moisture measurements were performed using a newly developed wetness tester (L-SALIVO), in which the wet portion was measured after 10 s. Further, clinical diagnosis of dry mouth was carried out using a clinical classification scale of the tongue mucosa (grade range, 0-3). RESULTS: It was possible to measure tongue dorsum moisture in all subjects with the wetness tester. The average moisture value was 0.1+/-0.2 mm in elderly subjects with a dry mouth grade of 2 (n = 8) or 3 (n = 12), while the average moisture value in the control subjects was 3.67+/-1.75 mm with a dry mouth grade of 0 (n = 4) or 1 (n = 2). Tester values and cliniical classification showed a positive co-relationship (r = 0.31, p < 0.05). CONCLUSIONS: Our results show that this new tester could be useful for evaluating oral dryness and diagnosing dry mouth.