Isolation and structural analysis of efficient autonomously replicating sequences (ARSs) of the yeast Candida utilis

The industrially important yeast Candida utilis is widely used in production of food and medical materials, but its host-vector system has not been well developed. We screened for compact and efficient ARSs to construct practically useful vectors. The C. utilis strain AHU3053 was found to be efficiently transformed by the conventional lithium acetate method and was used as the host. The C. utilis IAM4264 genomic library was constructed by inserting the partial Sau3AI digests in pRI51, which has a kanMX gene expressible in C. utilis. By examining 98 C. utilis G418-resistant transformants, five plasmids had the highest ARS activity. By trimming of the inserts, the 1490 and 552 bp fragments with transformation activity of over 10(3)/microg DNA were obtained from ARS3 and ARS4, respectively. Although several sequences identical to S. cerevisiae ARS consensus sequences (ACSs) were found in ARS3 and ARS4, our deletion analysis indicated that these were not essential for the activity. Because the minimal functional ARS fragment was also several-fold larger than that of S. cerevisiae, the C. utilis ARSs have some unique characteristics resembling the Sz. pombe ARSs. These ARSs were functional in other C. utilis strains tested and useful for constructing practical vectors.     

Strengthening of Preloaded RC Beams Using Hybrid Carbon Sheets

Reinforced concrete (RC) beams subject to service loads of 40 or 60% of steel yielding were strengthened using hybrid continuous carbon fiber sheets. The hybrid systems were made of high-strength and high-modulus carbon sheets, and compared with systems using only high-strength carbon. It was found that the use of high-modulus carbon sheets in hybrid systems could increase the yielding load, the flexural stiffness, the postyielding ductility, and reduce the crack opening in concrete. The slope changes on load-deflection curves at steel yielding are not noticeable in hybrid systems. The tensile strains developed in hybrid sheets after the fracture of high-modulus carbon are higher in magnitude and distributed in a larger area, leading to an ultimate carbon fracture with concrete crushing. These unique features are attributed to the high stiffness and low ultimate tensile strain of the high-modulus carbon fibers which stiffen the structures, avoid or delay the fiber-reinforced polymer debonding, and facilitate the deformability during their subsequent breakdown.     

Structure determination of tubular crystals of membrane proteins. IV. Distortion correction and its combined application with real-space averaging and solvent flattening

A method for correction of three-dimensional distortions has been developed for helical assemblies and applied to tubular crystals of Ca2+-ATPase. This method approximates distorted helical particles with short straight segments of different orientation parameters, which are determined by fitting them to the reference data in reciprocal space. Thus, the method follows Beroukhim and Unwin [Ultramicroscopy 70 (1997) 57], but is more extended to achieve better alignment and to cope with images of poor S/N ratio. Substantial improvements were achieved by dividing the reference image into the segments of optimal lengths in exactly the same way as the test, and treating the distortions in the near and far sides of a helical particle separately. The improvement was further enhanced when combined with real-space averaging [Yonekura, Toyoshima, Ultramicroscopy 84 (2000) 15] and solvent flattening [Yonekura, Toyoshima, Ultramicroscopy 84 (2000) 29], and most pronounced when all these three were applied iteratively.     

Expression of endothelial and inducible nitric oxide synthase in human glomerulonephritis

The presence of nitric oxide (NO) in the kidney has been implicated in the pathogenesis of human glomerulonephritis. However, the exact type of glomerular cells that express NO synthase (NOS) and the NOS isoform involved in the local production of NO has not been identified in the human diseased kidney. We examined the expression of three isoforms of NOS, inducible NOS (iNOS), endothelial NOS (eNOS) and brain NOS (bNOS) in the renal tissue of patients with IgA nephropathy (IgAN, N = 10), lupus nephritis (LN, N = 5), membranous nephropathy (MN, N = 5) and minimal change nephrotic syndrome (MCNS, N = 5). Sections were immunostained and the correlation between the expression of each NOS and the degree of glomerular injury in that section was also examined. Normal portions of surgically resected kidneys served as controls. eNOS was present in glomerular endothelial cells and endothelium of cortical vessels in the control and diseased kidneys. iNOS was localized in mesangial cells, glomerular epithelial cells and infiltrating cells in the diseased glomeruli, whereas immunostaining for iNOS was hardly detected in control kidneys. In addition, the expression pattern of eNOS in each glomerulus was the reverse of that of iNOS. In IgAN and LN, the extent of staining for eNOS correlated negatively with the degree of glomerular injury, while the extent of staining for iNOS correlated positively with the degree of glomerular injury in the same tissues. bNOS was not detected in normal or nephritic glomeruli. Our results indicate the presence of a NO pathway in human diseased kidney, and suggest that NO derived from eNOS and iNOS may be involved in the progression of renal diseases and that NO derived from each NOS may play an important role in different way in human inflamed glomeruli.     

Iodine-123-BMIPP myocardial washout and cardiac work during exercise in normal and ischemic hearts

Due to the changes in the frequency of penicillin-resistant strains of S. pneumoniae, it is necessary to perform surveillance studies of bacterial resistance. Isolates from the upper respiratory tract of asymptomatic children have been useful. There is no information about the difference between isolates from children with and without upper respiratory tract infection (URTI). The objective of the authors in this paper is to establish the prevalence of carrier-state, serotype and antimicrobial resistance of S. pneumoniae isolates from children with and without acute upper respiratory tract infection (URTI) in a rural area in Mexico. A cross-sectional comparative study was performed in Tlaxcala, Mexico. Children from one month 5 years of age were included. Nasopharyngeal swabs were obtained. Identification was done by international microbiology standards. Serotyping was done by the capsular Quellung test. The susceptibility testing was performed by the agar dilution method. Four-hundred and fifty patients were included. S. pneumoniae was isolated in 134 children (29.7%). Frequency of carriers was greater in patients with URTI (107/323) than without URTI (27/127) (33.1% vs. 21.1% p = 0.012, OR 1.84, IC 95% 1.1-3.08). The six most frequent serotypes were: 6B (16.4%); 19F (11.9%); 19A (6.7%); 14, 23F, and 35 (5.2% each), with no difference among the groups. Only 3% of the strains had high level resistance to penicillin, and 12.6% had intermediate resistance, and for ampicillin 4%, amoxicillin 4%, amoxicillin-clavulanate 4%, ceftriaxone 3%, cefotaxime 1.5%, erythromycin 6%, miocamycin 3%, chloramphenicol 4%, and vancomycin 0%. Trimethoprim-sulfamethoxazole resistance was very high (42%). In conclusion, colonization is higher in children with URTI. Five of the most frequent serotypes identified in this study were the same as those identified in patients with S. pneumoniae invasive diseases in Mexico City. In Tlaxcala, Mexico, beta-lactams could be the drug of choice for the treatment of S. pneumoniae lower respiratory tract infections. It is necessary to perform clinical assays to evaluate the efficacy of trimethoprim-sulfamethoxazole due to the high resistance in vitro.     

The secondary buckling and post-secondary-buckling behaviours of rectangular plates

Secondary buckling and post-secondary-buckling behaviours are theoretically studied for simply-supported rectangular plates, whose primary buckling modes of deflection contain more than half-waves in the load acting direction. Modal coupling effects more complex than one two-term-coupling effects are incorporated into the secondary buckling and post-secondary-buckling analyses. Then, unstable or stable symmetric secondary branching points are found on the post-primary-buckling paths and “snap through” motions involving an abrupt change in wave-form are shown to be possible. Wave-form variations along post-secondary-buckling paths are also disclosed by means of a numerical analysis of equilibrium paths.     

Threshold Stress for Crack Healing of Mullite Reinforced by SiC Whiskers and SiC Particles and Resultant Fatigue Strength at the Healing Temperature

A mullite/SiC whisker/SiC particle multi-composite, having excellent crack-healing ability and mechanical properties, was hot pressed in order to investigate the crack-healing behavior under stress and the resultant fatigue strength at the temperature of healing. A semi-elliptical surface crack 100 μm in surface length was introduced on each specimen. The pre-cracked specimens were crack healed under cyclic or constant stress by using a three-point bending stress at 1473 K, and the resultant bending strength and cyclic fatigue strength were measured at 1473 K. The pre-crack on the surface of the specimens could be healed even under stress. The threshold stresses for crack healing, as determined by evaluating the strengths of crack-healed specimens at a healing temperature of 1473 K, were 170 MPa for both constant and cyclic stresses, corresponding to 77% of the bending strength of the pre-cracked specimens. The static and cyclic fatigue behaviors of crack-healed specimens were also investigated at a healing temperature of 1473 K.     

Production of finely ground natural graphite particles with high electrical conductivity by controlling the grinding atmosphere

Natural graphite particles with high crystallinity sieved to obtain a particle size range of under 63 μm were ground with a ball mill, under various well-controlled grinding atmospheres such as N₂, O₂, He, H₂, and vacuum. The ratio, X_dif50/X_st50, i.e. between the 50 wt.% Stokes diameter and the 50 wt.% laser diffraction diameter, of the ground particles, was used as an index of the flakiness of the particles. The specific resistance of films composed of the ground graphite particles was systematically measured. The rate of reduction in the size of the particles by grinding was slow under an O₂-rich atmosphere such as 100% O₂ and dry air. On the other hand, it was relatively fast in vacuum, or under an N₂ or He atmosphere, and a gas mixture of 99% N₂ and 1% O₂. The rate of size reduction by grinding under a H₂ atmosphere was intermediate. In our experimental conditions, the flakiness of the ground particles increased with the decrease in the particles’ sizes. The electrical conductivity of the ground particles, however, tended to decrease with the decrease in their sizes. Under the condition that the Stokes diameter of the ground particles remains constant, the electrical conductivity of films made from the ground particles increases with the increase in the flakiness of the particles. It was finally determined from our systematic grinding experiments that small flaky particles, which had a size, X_st of ∼1 μm, with a high electrical conductivity can be produced by grinding in a gas mixture of 99% N₂ and 1% O₂. In this case, the flaky shape of the ground particles was visually confirmed by scanning electron microscopy.     

Densification and Thermal Conductivity of AIN Doped with Y2O3, CaO, and Li2O

Small amounts of Li₂O result in sintering in the AIN-Y₂O₃-CaO and AIN-CaO systems at firing temperatures <1600°C. The effect is ascribed to reduction of the liquidus temperature. Furthermore, Li₂O is removed by volatization at temperatures from 1300° to 1600°C, and its content decreases several ppm from the initial 0.3 wt%. Li₂O-doped AIN specimens containing Y₂O₃ and CaO additives are well densified by firing at 1600°C for 6 h, and their thermal conductivity is 135 W^.m^−1.K⁻¹.The effect of Li₂O addition on sintering and thermal conductivity also is discussed through thermo-dynamic considerations.     

High Thermal Conductivity in Silicon Nitride with Anisotropie Microstructure

Silicon nitride was fabricated by tape casting of α-Si₃N₄ powder with 5 wt% Y₂O₃ and 5 vol% rodlike β-Si₃N₄ seed particles, followed by tape stacking, hot pressing under 40 MPa, and annealing at 1850°C for 2-66 h under a nitrogen pressure of 0.9 MPa. Silicon nitrides fabricated by this procedure exhibited a highly anisotropic microstructure with large elongated grains (developed from seed particles) uniaxially oriented parallel to the casting direction. Thermal conductivities parallel to the grain alignment were much higher than those measured in other directions and exhibited high values of up to 120 W/(m.K). The anisotropic thermal conductivity of the specimen could be explained by the rule of mixture, considering that large elongated grains developed from seeds have higher thermal conductivity than a small-grained matrix.