Postoperative decrease of regional volumetric bone mineral density measured by quantitative computed tomography after lumbar fusion surgery in adjacent vertebrae.

We investigated the effect of posterior lumbar fusion surgery on the regional volumetric bone mineral density (vBMD) measured by quantitative computed tomography. Surgery negatively affected the regional vBMD in adjacent levels. Interbody fusion was independently associated with vBMD decline and preoperative epidural steroid injections (ESIs) were associated with less postoperative vBMD decline. INTRODUCTION: Few studies investigate postoperative BMD changes after lumbar fusion surgery utilizing quantitative computed tomography (QCT). Additionally, it remains unclear what preoperative and operative factors contribute to postoperative BMD changes. The purpose of this study is to investigate the effect of lumbar fusion surgery on regional volumetric bone mineral density (vBMD) in adjacent vertebrae and to identify potential modifiers for postoperative BMD change. METHODS: The data of patients undergoing posterior lumbar fusion with available pre- and postoperative CTs were reviewed. The postoperative changes in vBMD in the vertebrae one or two levels above the upper instrumented vertebra (UIV+1, UIV+2) and one level below the lower instrumented vertebra (LIV+1) were analyzed. As potential contributing factors, history of ESI, and the presence of interbody fusion, as well as various demographic/surgical factors, were included. RESULTS: A total of 90 patients were included in the study analysis. Mean age (±SD) was 62.1 ± 11.7. Volumetric BMD (±SD) in UIV+1 was 115.4 ± 36.9 mg/cm3 preoperatively. The percent vBMD change in UIV+1 was - 10.5 ± 12.9% (p < 0.001). UIV+2 and LIV+1 vBMD changes showed similar trends. After adjusting with the interval between surgery and the secondary CT, non-Caucasian race, ESI, and interbody fusion were independent contributors to postoperative BMD change in UIV+1. CONCLUSIONS: Posterior lumbar fusion surgery negatively affected the regional vBMDs in adjacent levels. Interbody fusion was independently associated with vBMD decline. Preoperative ESIs were associated with less postoperative vBMD decline, which was most likely a result of a preoperative decrease in vBMD due to ESIs.

Effect of sodium conductance variations on electrical behavior of a neocortical neuron model.

Neocortical pyramidal neurons are capable of intrinsic regenerative firing. A mathematical model introduced by Delord et al. [2] of these neurons based on the Hodgkin-Huxley formalism, varied the values of two parameters of the model, i.e. the maximal persistent sodium conductance (gNaP) and the leakage one (gl), and revealed the (gNaP, gl) parameter space supporting regenerative firing. The present study focused on another parameter of this model, i.e. the maximal fast sodium conductance (gNa), to investigate the (gNaP, gNa) parameter space involved in regenerative firing. When gNa was completely blocked, regenerative firing was suppressed. In addition, the gNa threshold necessary to induce regenerative firing was almost independent of the gNaP level. Finally, our results were compared with those of other types of neurons.

Novel types of bistability in a model of a bursting pacemaker neuron RPa1 from the snail, Helix pomatia.

The RPa1 neuron identified in the snail, Helix pomatia, produced a variety of electrical activities (e.g. bursting and spiking). A previously developed mathematical model, which described these activities, revealed bistability between bursting and chaotic spiking, where chaotic spiking was transformed into bursting by a short-lasting external stimulus, and vice versa. The present study used this model to detect other types of bistability, i.e. bistability between bursting and period-2 spiking and between bursting and period-4 spiking (period-2 and -4 spiking are generated by period-doubling bifurcation). This contributes to our understanding of the electrophysiological properties of RPa1.

Analysis of the electrosensory pyramidal cell bursting model for weakly electric fish: model prediction under low levels of dendritic potassium conductance.

Pyramidal cells in the electrosensory lateral line lobe (ELL) of weakly electric fish produce burst discharge. A Hodgkin-Huxley-type model, called ghostburster, consisting of two compartments (soma and dendrite) reproduces ELL pyramidal cell bursting observed in vitro. A previous study analyzed the ghostburster by treating Is and gDr,d as bifurcation parameters (Is: current injected into the somatic compartment and gDr,d: maximal conductance of the delayed rectifying potassium current in the dendritic compartment) and indicated that when both Is and gDr,d are set at particular values, the ghostburster shows a codimension-two bifurcation at which both saddle-node bifurcation of fixed points and saddle-node bifurcation of limit cycles occur simultaneously. In the present study, the ghostburster was investigated to clarify the bursting that occurred at gDr,d values smaller than that at the codimension-two bifurcation. Based on the number of spikes per burst, various burst patterns were observed depending on the (Is, gDr,d) values. Depending on the (Is, gDr,d) values, the burst trajectory in a phase space of the ghostburster showed either a high or a low degree of periodicity. Compared to the previous study, the present findings contribute to a more detailed understanding of ghostburster bursting.

Probing the metal-insulator phase transition in the (DMEDO-EBDT)2PF6 single crystal by optical measurements.

The temperature and polarization dependence of the optical reflectivity  spectra of a quasi-one-dimensional 1/4-filled band system, (DMEDO-EBDT)(2)PF(6), have been investigated. We observed clear anisotropy in the electronic structures corresponding to the anisotropic transport properties. The appearance of a charge gap (E(g) > 0.1 eV) and transfer of the spectral weight accompanied by the metal-insulator phase transition were clearly observed. In addition, a split of the intramolecular vibrational modes was observed, which strongly suggested the existence of charge disproportionation in the low temperature phase. We also observed a photoinduced reflectivity change, which implied the occurrence of a photoinduced phase transition from the low temperature insulating phase to the high temperature metallic phase.

Evaluation of a new fiber-grating vision sensor for assessing pulmonary functions in healthy and COPD subjects.

Spirometry is practically the only tool to evaluate pulmonary functions. Other automatic systems comparable to spirometry are expected. A fiber-grating (FG) vision sensor is a non-contact respiratory monitoring system to detect changes in volumes by measuring the movement of laser spots on the body surface. We examined the contributions of the FG sensor to evaluating pulmonary functions. The FG sensor showed a linear correlation with spirometry in tidal volumes (TV) obtained from five controls (R = 0.98, P < 0.0001). We also showed agreement of TV between the two devices using Bland-Altman analysis. TV measured by the FG sensor were reproducible and applicable to distinct subjects. To detect airway obstruction, we performed forced expiration in controls (n = 16) and chronic obstructive pulmonary disease (COPD) patients (n = 18) with the FG sensor and spirometry. Forced expiratory volume in 1 s (FEV(1)) and FEV(1)/forced vital capacity in COPD patients were lower than those in controls by the FG sensor. In addition, prolonged expiration in natural breathing by the FG sensor was related to airflow limitation by spirometry. The FG sensor was helpful to measure volume changes and to evaluate pulmonary functions in controls and patients with COPD. Its upcoming clinical applications are promising for simplicity and feasibility.

The effect of variations in sodium conductances on pacemaking in a dopaminergic retinal neuron model.

Dopaminergic neurons in the retina show spontaneous tetrodotoxin-sensitive pacemaking, which has been explained by a reduced Hodgkin-Huxley-type computer model. The present study used this model to investigate the effect of variations in transient and persistent sodium conductance values on pacemaking, under variable leakage conductance levels. This study indicated that transient sodium conductance plays an indispensable role in pacemaking, which occurs under conditions in which only a persistent sodium conductance is considerably reduced, thus contributing to a detailed understanding of the relationship between sodium conductance and pacemaking.

Protective immunity of SpaA-antigen producing Lactococcus lactis against Erysipelothrix rhusiopathiae infection.

AIMS: To develop an economical, safe and simple vaccination system against swine erysipelas using SpaA-antigen producing Lactococcus lactis. METHODS AND RESULTS: The spaA gene of Erysipelothrix rhusiopathiae was inserted into a shuttle plasmid pSECE1 to construct pSECE1.3. The SpaA produced in L. lactis maintained a stable antigenicity without degrading in growth. After mice were inoculated intranasally and orally with pSECE1.3-carrying L. lactis cells, IgG and IgA specific to SpaA were detected, and all the mice survived a challenge with 100 LD(50) of E. rhusiopathiae Tama-96 in the inner thigh. CONCLUSIONS: SpaA-producing L. lactis appears useful as an effective subunit vaccine against swine erysipelas. SIGNIFICANCE AND IMPACT OF THE STUDY: In this vaccination system, purification of the antigen and injection are unnecessary, leading to a reduced production cost, reduced labour and less stress to the animals. This vaccination system of the lactic acid bacteria should be a safe and suitable vehicle for a polyvalent vaccine.

Phylogenetic diversity and similarity of active sites of Shiga toxin (stx) in Shiga toxin-producing Escherichia coli (STEC) isolates from humans and animals.

Nucleotide sequences of Shiga toxin (Stx) genes in STEC from various origins were determined and characterized by phylogenetic analysis based on Shiga toxin (Stx) with those deposited in GenBank. The phylogenetic trees placed Stx1 and Stx2 into two and five groups respectively, and indicated that Stx1 in sheep-origin STEC were placed into a different group from those in other STEC, and that Stx2 of deer-origin STEC also belonged to the unique group and appeared to be distantly related to human-origin STEC. On the other hand, Stx of STEC isolated from cattle, seagulls and flies were closely related to those of human-origin STEC. Such a diversity of Stx suggested that STEC might be widely disseminated in many animal species, and be dependent on their host species or their habitat. In addition, the active sites in both toxins were compared; the active sites in both subunits of Stx in all the animal-origin STEC were identical to those in human-origin STEC, suggesting that all the toxin of STEC from animals might be also cytotoxic, and therefore, such animal-origin STEC might have potential pathogenicity for humans.

A simple and sensitive detection system for Bacillus anthracis in meat and tissue.

AIMS: To detect and isolate Bacillus anthracis from meat and tissue by rapid and simple procedures. METHODS AND RESULTS: Bacillus anthracis Pasteur II cells were added to 1 g lymph node and pig meat, which were then cut into small pieces and suspended in PBS. Aliquots were spread on Bacillus cereus selective agar (BCA) plates to isolate B. anthracis cells, and incubated in trypticase soy broth. The enrichment culture was used for nested PCR with B. anthracis specific primers, which were to confirm the presence of B. anthracis chromosomal DNA and the pXO1/pXO2 plasmids. CONCLUSION: One cell of B. anthracis was detected by nested PCR from 1 g of the samples, and was also isolated on BCA plates according to colony morphology within two days. SIGNIFICANCE AND IMPACT OF THE STUDY: These results could be useful for detecting animals with latent anthrax, and meat contaminated with B. anthracis, rapidly and simply.

Genetically modified Shiga toxin 2e (Stx2e) producing Escherichia coli is a vaccine candidate for porcine edema disease.

Porcine edema disease (ED) is an enterotoxaemia in pigs after weaning, caused by Shiga toxin 2e (Stx2e) producing Escherichia coli. Recently in Japan, outbreaks of ED are re-emerging in pig production. In this study we constructed a mutant that retained immunogenicity but lost Vero cell cytotoxicity, which produced genetically modified toxin (Stx2e*) by replacing glutamate with glutamine at position 167 and arginine with leucine at position 170 of the A subunit. The stx(2e)* gene was replaced with the stx(2e)gene of the wild type virulent strain by homologous recombination. As the parent wild strain was pathogenic to pigs but the mutant was not, the mutant named as YT106 was given to the pigs to examine its protective immunity against ED. All 20 pigs vaccinated with YT106 survived, but only eight of the 20 non-vaccinated pigs survived after the challenge with a wild strain. Also, the eight pigs that survived had decreased rates of gain relative to those of the controls. Blood IgG and intestinal IgA titres increased 3.3 and 1.6 times more than the control, respectively, showing that YT106 might be a good candidate of a live attenuated vaccine strain to protect against ED.

Effects of rearing conditions on the colonization of Salmonella enteritidis in the cecum of chicks.

Salmonella enteritidis is the cause of human salmonellosis associated with contaminated eggs. In this study, we artificially challenged S. enteritidis to chicks just after hatching, and the effects of breeding conditions on the intestinal carriage of S. enteritidis were examined. S. enteritidis was not directly detected from spleen, liver and blood, but were constantly isolated from the cecal contents throughout the experiment. When chicks were reared in the unsanitary conditions and in the high housing density, the numbers of S. enteritidis increased. The subsequent experiment was undertaken to examine whether the antibacterial additive in a feed would have any impact on S. enteritidis colonization in chicks. Some antibiotic effective on the growth promotion had an influence on S. enteritidis colonization.

Does enterohemorrhagic Escherichia coli O157:H7 enter the viable but nonculturable state in salted salmon roe?

An outbreak caused by salted salmon roe contaminated with enterohemorrhagic Escherichia coli O157 occurred in Japan in 1998. Since about 0.75 to 1.5 viable cells were estimated to cause infection, we presumed that O157 might enter the viable but nonculturable (VNC) state in salted salmon roe and consequently that viable cell numbers might be underestimated. Although patient-originating O157 cells could not grow on agar plates after 72 h of incubation in 13% NaCl, they were resuscitated in yeast extract broth, and more than 90% of the cells were shown to be viable by fluorescent staining, suggesting that almost all of them could enter the VNC state in NaCl water. Roe-originating O157 was resistant to NaCl because it could grow on agar after 72 h of incubation in NaCl water, but about 20% of cells appeared to enter the VNC state. Therefore, germfree mice were infected with O157 to examine the resuscitation of cells in the VNC state and the retention of pathogenicity. O157 that originated in roe, but not patients, killed mice and was isolated from the intestine. However, these isolates had become sensitive to NaCl. O157 cells of roe origin incubated in normal media also killed mice and were isolated from the intestine, but they also became transiently NaCl sensitive. We therefore propose that bacterial cells might enter the VNC state under conditions of stress, such as those encountered in vivo or in high salt concentrations, and then revive when those conditions have eased. If so, the VNC state in food is potentially dangerous from a public health viewpoint and may have to be considered at the time of food inspection. Finally, the establishment of a simple recovery system for VNC cells should be established.

Detection and characterization of Shiga toxin-producing Escherichia coli from seagulls.

Shiga toxin (Stx)-producing Escherichia coli (STEC) strains isolated from a seagull in Japan were examined. A total of 50 faecal samples was collected on a harbour bank in Hokkaido, Japan, in July 1998. Two different STEC strains, whose serotypes were O136:H16 and O153:H-, were isolated from the same individual by PCR screening; both of them were confirmed by ELISA and Vero cell cytotoxicity assay to be producing active Stx2 and Stx1, respectively. They harboured large plasmids, but did not carry the haemolysin or eaeA genes of STEC O157:H7. Based on their plasmid profiles, antibiotic resistance patterns, pulsed-field gel electrophoresis analysis (PFGE), and the stx genes sequences, the isolates were different. Phylogenic analysis of the deduced Stx amino acid sequences demonstrated that the Stx toxins of seagull-origin STEC were closely associated with those of the human-origin, but not those of other animal-origin STEC. In addition, Stx2phi-K7 phage purified from O136 STEC resembled Stx2phi-II from human-origin O157:H7, and was able to convert non-toxigenic E. coli to STEC. These results suggest that birds may be one of the important carriers in terms of the distribution of STEC.

Antibacterial activity of chaff vinegar and its practical application.

Since enterohemorrhagic Escherichia coli O157, Salmonella, etc., sometimes contaminate animal feces and may cause infectious diseases to humans, it is important to remove pathogenic bacteria from domestic animal waste. For the purpose, we examined the antibacterial activity of chaff vinegar. We found that the chaff vinegar inhibited the growth of pathogenic bacteria immediately in vitro but not efficiently spores and lactic acid bacteria. Further, it removes bacteria, especially Enterobacteriaceae, from animal feces and the surface of the concrete-floor in the cattle barn. Chaff vinegar is advertised as a natural chemical substance for a soil conditioner, to promote the composting and to deodorize their smell. Chaff vinegar may be useful for organic agriculture without enteric pathogenic bacteria.

Surface antigen, SpaA, of erysipelothrix rhusiopathiae binds to Gram-positive bacterial cell surfaces.

In a previous study, we isolated the spaA gene encoding the surface protective antigen A, SpaA, of Erysipelothrix rhusiopathiae, and found that the N-terminal region of SpaA was responsible for protective immunity against erysipelas and that the C-terminal region contained eight repeat units consisting of 20 amino acids comprising the binding domain on the Erysipelothrix cell surface. In this study, using recombinant SpaA proteins, we showed that the repeat region bound to the cell surfaces of various Gram-positive bacterial cells, SpaA was a membrane-associated protein, this association depended on the interaction with choline residues in teichoic acid, and SpaA bound to lipoteichoic acid (LTA) of Bacillus subtilis and Staphylococcus aureus. These results showed that LTA was required for the surface association of SpaA in E. rhusiopathiae and that such an association might be common among Gram-positive bacterial cells. We suggested that an LTA-SpaA complex might have an important role in the E. rhusiopathiae infection process.

Tumor necrosis factor alpha and gamma interferon are required for the development of protective immunity to secondary Corynebacterium pseudotuberculosis infection in mice.

The production and role of endogenous cytokines during the course of secondary Corynebacterium (C.) pseudotuberculosis infection were investigated in mice. When immunized mice were challenged on day 28 after primary infection, tumor necrosis factor alpha (TNF-alpha) and interferon gamma (IFN-gamma) were found to appear at 3 hr and to reach the maximum at 24 hr after challenge. Spleen cells of mice primarily infected from 2 to 8 weeks before produced a significant amount of TNF-alpha and IFN-gamma when stimulated with formalin-killed bacteria. However, they could not produce detectable amounts of IL-4. The administration of anti-TNF-alpha monoclonal antibody (MAb) and IFN-gamma MAb increased bacterial proliferation in the organs of immune mice and exacerbated the secondary infection. Injection of anti-CD4 MAb alone or anti-CD4 plus anti-CD8 MAbs resulted in significantly increased mortality and a marked suppression of bacterial elimination as well as cytokine production of secondarily infected mice, while the treatment with anti-CD8 MAb alone showed no effect on either the resistance or cytokine production of mice. These results suggest that CD4, probably Th1 T cells, play an important role for establishment of protective immunity against secondary C. pseudotuberculosis infection by secreting TNF-alpha and IFN-gamma.

Establishment of the PCR system specific to Salmonella spp. and its application for the inspection of food and fecal samples.

We established the PCR detection system specific to Salmonella species using Salmonella enterotoxin gene (stn). The detection limit was one bacterial cell per one gram of fecal and minced-meat samples using enrichment procedure by Tripticase soy broth or Salmonella enrichment broth, respectively. We concluded that this PCR system is useful for the practical application in the field of the public hygiene.

Molecular epidemiological study on tetracycline resistance R plasmids in enterohaemorrhagic Escherichia coli O157:H7.

Restriction patterns obtained with EcoRI and Southern hybridization were used for the differentiation of tetracycline-resistant (Tet(r)) R plasmids in enterobaemorrhagic Escherichia coli (EHEC) O157:H7 isolates from a mass outbreak at a kindergarten in Obihiro-City, Hokkaido, Japan, 1996. Two kinds of Tet(r) R plasmids of 50 and 95 kb were detected. The 50-kb plasmids were identical to each other, while the 93-kb plasmids were of three types that were very similar to each other. The tet genes of both 50- and 95-kb R plasmids were 100% identical to the tet gene of pSC101 and all plasmids hybridized to a probe for tet. Because food-origin O157 strains were sensitive to tetracycline, we concluded that such Tet(r) R-plasmids might transfer to drug-sensitive O157 strains in the infected individuals.