Performance of a rapid human metapneumovirus antigen test during an outbreak in a long-term care facility.

Using a newly developed rapid test, an outbreak of human metapneumovirus (HMPV) infection in a long-term care facility was detected within only 2 days after the onset of symptoms in a putative index case. The outbreak was almost under control within 8 days mainly by zoning patients, with the exception of two cases of HMPV that were diagnosed 16 and 17 days after the onset of the outbreak. According to an immunological diagnosis as well as the rapid test, it was eventually proven that 18 patients had HMPV infections. We suspected that even asymptomatic residents, who had not been completely separated from the facility population, were a source of infection. That suggested that all asymptomatic residents should be tested and that the separation of the infected patients should be absolute, if an outbreak of HMPV infection is suspected in such a facility.

Coexistence effect of UVA absorbers to increase their solubility and stability of supersaturation.

OBJECTIVE: Sunscreens containing UVA absorbers in high concentrations are expected to be developed, since recent studies have suggested the possibility of involvement of UVA ray in skin cancer and early skin aging. Solubility and stability of supersaturation of UVA absorbers in UVB absorber were determined in the absence and the presence of cosmetic oil. Coexistence effect of UVA absorbers was analyzed to dissolve them in high concentrations. METHODS: Two UVA absorbers, diethylamino hydroxybenzoyl hexyl benzoate (DHHB) and butyl methoxydibenzoylmethane (BMDM), a UVB absorber, 2-ethylhexyl methoxycinnamate (EHMC), and a cosmetic oil, 2-ethylhexyl ester of oligomer of hydroxystearic acid (EH-O-HSA), were used. Their solutions were prepared at 80°C and cooled to 5°C. The solid DHHB and/or BMDM were added to it, and the time evolution of concentrations of the UVA absorbers in the solution phase was monitored. RESULTS: At the saturation in the absence of EH-O-HSA at 5°C, weight ratio of DHHB and BMDM to EHMC was 0.39/1.00 and 0.22/1.00, respectively. Addition of EH-O-HSA slightly changed the solubility of DHHB and BMDM. When the weight ratio of EH-O-HSA to EHMC was 0.20/1.00, weight ratio of DHHB and BMDM to EHMC was 0.35/1.00 and 0.25/1.00, respectively at the saturation at 5°C. In the presence of EH-O-HSA, a strong coexistence effect of DHHB and BMDM was found on their solubility. A thermodynamically stable saturated solution at 5°C having the composition that DHHB: BMDM: EHMC: EH-O-HSA = 0.47: 0.46: 1.00: 0.20 was obtained by the simultaneous addition of solid DHHB and BMDM into the initial solution. CONCLUSION: The solution type composite having the highest concentrations of DHHB and BMDM prepared in this study exhibited critical wavelength at 368 nm that was just below the border for sunscreens being qualified as 'Broad Spectrum' protection under the new rule launched by US FDA.

Abortion in a thoroughbred mare associated with an infection with avirulent Rhodococcus equi.

An eight-year-old thoroughbred mare with no previous history of illness aborted a fetus at 196 days of gestation, and its internal tissues were examined immunohistologically and bacteriologically. The placenta was not examined, but specimens of the intrauterine fluids and the dam's faeces were collected four days after the abortion and examined bacteriologically. No significant histological lesions were found in the fetus but the amnion and the umbilical cord were oedematous and had petechial haemorrhages. Rhodococcus equi was isolated in pure culture from the lung, heart and stomach contents of the fetus and from an intrauterine specimen and faeces of the dam. The anti-R equi antibody titre of the mare was high after the abortion. The diagnosis was confirmed in the lung of the fetus by immunohistochemical staining with R equi-specific antibodies. Isolates from the fetus and mare were identified as avirulent R equi by pcr and the mouse pathogenicity test. The avirulent isolates were characterised by pulsed-field gel electrophoresis, which yielded only one VspI profile in all the isolates from the fetus and its dam.

Current-free double-layer formation in inductively coupled plasma in a uniform magnetic field.

The axial profiles of plasma parameters for low and moderate pressures, such as the plasma potential, electron temperature, and number density, have been evaluated in magnetized inductively coupled plasma. The experimental results revealed in both cases the existence of a genuine current-free double-layer structure, separating two plasma regions with different properties. Based on the experimental results, a physical scenario for the self-assembling of the double layer is proposed. Also, the axial profile of the electron number density downstream is analyzed, emphasizing the role of neutral metastable ionization, and a simple analytical model is developed to fit the experimental data. The model allows the estimation of neutral metastable number density downstream and the recombination rate coefficient.

Effect of concanavalin A on the activity of membrane-bound and detergent-solubilized Mg2+ -ATPase.

Plasma membranes were isolated after binding liver and hepatoma cells to polylysine-coated polyacrylamide beads, and the effect of concanavalin A on the membrane-bound Mg2+ -ATPase and the Mg2+ -ATPase solubilized by octaethylene glycol monododecyl ether (C12E8) was studied. In the experiment of membrane-bound Mg2+ -ATPase, plasma membranes were pretreated with Concanavalin A and the activity was assayed. Concanavalin A stimulated the activity of both liver and hepatoma enzymes assayed above 20 degrees C. Concanavalin A abolished the negative temperature dependency characteristic of liver plasma membrane Mg2+ -ATPase. On the other hand, Concanavalin A prevented the rapid inactivation due to storage at -20 degrees C, which was characteristic of hepatoma plasma membrane Mg2+ -ATPase. With solubilized Mg2+ -ATPase from liver plasma membranes, the negative temperature dependency was not observed. Concanavalin A, which was added to the assay medium, stimulated the activity of the enzyme solubilized in C12E8 at a high ionic strength. However, Concanavalin A failed to show any effect on the enzyme solubilized in C12E8 at a low ionic strength. With solubilized Mg2+ -ATPase from hepatoma plasma membranes, Concanavalin A could not prevent the inactivation of the enzyme during incubation at -20 degrees C.

Ca2+- or Mg2+-dependent ATPase in plasma membrane of cultured endothelial cells from bovine carotid artery.

ATPase was found in plasma membrane of cultured endothelial cells from bovine carotid artery. The activity of the enzyme solubilized by octaethyleneglycol mono-n-dodecyl ether was enhanced by the addition of Ca2+ or Mg2+ and was not affected by F-actin and ouabain. Vmax was 2.8 and 10.0 mumol Pi/mg protein per h for Ca2+- and Mg2+-dependent activity, respectively, and the corresponding Km was 4.8 X 10(-4) M and 3.2 X 10(-4) M. Molecular weight of the protein was estimated to be approx. 250 000, as determined by activity-staining electrophoresis with polyacrylamide gels.

Antibody-scanning and epitope-tagging methods; molecular mapping of proteins using antibodies.

Because synthetic short peptides bearing critical binding residues, can chemically mimic the folded antigenic determinants on proteins, short synthetic peptides can generate antibodies that react with cognate sequences in intact folded proteins. According to this mimotope theory, we produced site-specific antibodies by immunization with short peptides which overlapped each other and covered the entire protein, and used them for domain mapping of influenza virus RNA polymerase (antibody-scanning method). We also used a tagged-epitope and its monoclonal antibodies for topology mapping of clathrin light chains in clathrin triskelions by electron microscopy. Both methods using specific epitopes in combination with their antibodies enable us to determine the domains of interesting proteins systematically without the need to generate monoclonal antibodies or mutant proteins.

Inhibition of influenza virus replication in cultured cells by RNA-cleaving DNA enzyme.

Influenza virus replication has been effectively inhibited by antisense phosphothioate oligonucleotides targeting the AUG initiation codon of PB2 mRNA. We designed RNA-cleaving DNA enzymes from 10-23 catalytic motif to target PB2-AUG initiation codon and measured their RNA-cleaving activity in vitro. Although the RNA-cleaving activity was not optimal under physiological conditions, DNA enzymes inhibited viral replication in cultured cells more effectively than antisense phosphothioate oligonucleotides. Our data indicated that DNA enzymes could be useful for the control of viral infection.

Japanese monozygotic twins with familial amyloidotic polyneuropathy (FAP) (ATTR Val30Met).

Twenty-nine-year-old twin brothers having the amyloidogenic transthyretin (ATTR) Val30Met gene developed the clinical symptoms of familial amyloidotic polyneuropathy (FAP) in 1995. The twins had the same educational background and lived in the same district. FAP manifestations were similar in both cases, although electromyographic examinations revealed sensorimotor polyneuropathy in No. 1 and sensory polyneuropathy in No. 2. DNA analysis revealed that they were monozygotic twins. In addition to environmental factors, genetic factors may play an important role in determining the onset of FAP.

Densitometric determination of human papillomavirus DNA quantities by chromato-scanning in the fluorescence mode.

The quantitation of human papillomavirus DNA isolated from warts by chromato-scanning (fluorescence mode) photographs of ethidium bromide-stained agarose gels is described. Excitation at 200 nm (with a cutoff filter at 400 nm) generates fluorescence from the white portion of the printing paper. The fluorescent intensity correlated with the quantities of DNA in the band of interest. The amounts of DNA were determined using calibration curves of approximately the same size as lambda phage DNA fragments. This general method of quantification is applicable to photographs of other types of polynucleotides capable of being separated and stained in a gel medium.

Synthesis and characterization of polymer-coated mixed-functional stationary phases with several different hydrophobic groups for direct analysis of biological samples by liquid chromatography.

Three types of polymer-coated mixed-functional (PCMF) silica column-packing materials for the direct analysis of biological liquids have been synthesized. These packing materials all have polyoxyethylene groups as their hydrophilic part but differ in their hydrophobic part (methyl groups for Me-POE, phenyl groups for Ph-POE, and octyl groups for Oc-POE). Retention characteristics with respect to several drug molecules, and protein recovery, were studied for each of the three types with different amounts of hydrophobic groups attached. Of the PCMFs prepared, Oc-POE showed the greatest overall retention, and Me-POE displayed the greatest protein recovery.

Synthesis of polymer-coated mixed-functional packing materials for direct analysis of drug-containing serum and plasma by high-performance liquid chromatography.

Silicone polymer-coated mixed-functional (PCMF) silica packing materials were developed for the direct determination of drugs contained in serum or plasma. The new stationary phases were prepared by the following three-step procedure: (1) coating porous silica with a silicone polymer; (2) partial introduction of hydrophobic groups; and (3) introduction of hydrophilic groups. Two PCMFs were synthesized, one having polyoxyethylene groups as hydrophilic groups (PCMF-POE) and the other having oligoglyceryl groups (PCMF-OG). PCMF-POE showed higher recoveries for injected proteins and a greater overall retention for drug molecules than PCMF-OG. PCMF-POE did not show any column deterioration in 500 serum sample injections (10 ml in total).

Analysis of ibuprofen metabolites by semi-microcolumn liquid chromatography with ultraviolet absorption and pulsed amperometric detectors.

Semi-microcolumn (1.5 mm i.d.) liquid chromatography (LC) system with ultraviolet (UV) and pulsed amperometric detector (PAD) was constructed for analysis of ibuprofen metabolites in human urine. PAD was connected in series with the UV detector, and an alkaline solution was post-column added to a mobile phase after the UV detector. By a gradient elution, five ibuprofen metabolites were detected with UV detection from 1 microliter of human urine at 3.5 h after the administration, and information concerning their glucuronation was simultaneously obtained by PAD response.

Unusual homicide by air gun with pellet embolisation.

A homicidal case by an air gun involving a 73-year-old woman who was found dead in her home is described. At autopsy, two gunshot wounds were observed on the left anterior chest and the right back. There was neither exit wound nor powder burns. The fatal pellet, 0.9 g and 5.5mm diameter, penetrated the left anterior chest and the heart to cause cardiac tamponade (200 ml) before lodging in the left subclavian artery. Death was due to cardiac tamponade. This case has exhibited two rare phenomena: homicide by an air gun and pellet embolisation. This is the first homicide case by use of air gun in Japan and demonstrates that although it may be rare, air guns can be used as potential homicidal weapons. This experience will alert our society and others on the potential hazards of air guns we have tended to regard as toys.

Effects of protein phosphorylation with p43v-abl and protein kinase A catalytic subunit on fluorescence intensity.

To study the effect of phosphorylation on protein conformation, a fluorescence spectroscopic study was performed on phosphorylated enolase and histoneH1 proteins. The peak of fluorescence was 330 and 360 nm for each protein, respectively, when excited at 287 nm. The intensities of the fluorescence were measured during the phosphorylation reactions with the protein kinase A and p43v-abl, for serine, threonine and tyrosine, respectively. Slightly increased intensities at 330 and 360 nm for enolase and histoneH1 protein were observed by phosphorylation with p43v-abl, whereas decreased intensities occurred with the protein kinase A catalytic subunit. These data suggest that micro-structural changes are induced at the residue, either tyrosine, serine or threonine in the protein.

Subcloning and trial of expression of the protein kinase catalytic domain of RSV-src gene.

The gene of catalytic domain of the protein kinase of RSV-scr was cloned into the BamHI cloning site of translation vector pET-8c which containing T7 RNA polymerase promotor, and transformed BL21 (DE3) pLys S (Studier and Moffatt, 1986). The putative molecular weight of the protein was about 33 kd as evaluated on the basis of its nucleotide size showed the identical mobility in SDS-polyacrylamide gel electrophoresis. However, yield of protein production was not high, probably, because of its instability in Escherichia coli.

Interrupted aortic arch with ventricular septal defect: an unsuspected fatal congenital anomaly in forensic pathology.

Interrupted aortic arch (IAA) is a relatively rare cardiovascular anomaly worldwide and is very rarely encountered in forensic pathology. A case of clinically unsuspected fatal IAA in a 5-day old neonate is described. A full term female baby was declared dead on arrival at an emergency hospital few hours after having been discharged from a maternity hospital following delivery. There were no noticeable signs and symptoms reported during the short postnatal period. Antenatal medical history was unremarkable and there were no abnormalities noticed at birth. Autopsy revealed an unsuspected IAA arch with ventricular septal defect (VSD) as cause of sudden unexpected death. Although amenable to surgical intervention, IAA may not produce obvious clinical symptoms as was the case for this baby. Such sudden unexpected deaths raise medico-legal questions as to the manner and the precise cause of death. The importance of a full forensic autopsy to clarify such a sudden death was re-emphasized.

Death as a result of heat stroke in a vehicle: an adult case in winter confirmed with reconstruction and animal experiments.

A 54-year-old man was found dead in the driver's seat of his vehicle on a winter's day. Investigations of the vehicle revealed that the engine was running, and the car heater was left on with the maximum temperature and velocity. The body was found excessively sweating. Rectal temperature of the body was 43 degrees C at 10 h post mortem. In autopsy, several superficial skin burns were observed on the face, the shoulders and the legs. The lungs were heavily congested and hemorrhagic. The liver showed typical alcohol-induced micronodular cirrhosis. The alcohol concentrations were 0.17% in the blood of both the left and the right heart, 0.17% in the femoral-vein blood, 0.21% in the bladder urine and 0.34% in the gastric contents. A reconstruction experiment demonstrated that the temperature inside the vehicle rose rapidly and reached 50-58 degrees C in 3 h. Animal experiments showed that the temperature threshold for rats to succumb to heat was between 40 and 45 degrees C. This case shows that heat stroke in a vehicle can occur in adults with chronic diseases or alcoholism, such as in this particular case, even in the winter.