Effect of High Hydrostatic Pressure Treatment Combined with Alkaline Electrolyzed Water on the Injury and Growth Characteristics of Bacterial Spores.

The characteristics of growth and injury of Bacillus subtilis spores whose heat resistance was reduced by the treatment of high hydrostatic pressure processing (HPP) combined with/without alkaline electrolyzed water (AlEW) were investigated. The delay in the lag phase of growth curve observed when used in combination with AlEW, especially at 50 MPa or less, and was prolonged by about 4 to 6 hours. However, the effects of temperature and time during treatment on the lag phase were not significantly different between solo-HPP and in combination with AlEW. The injury characteristics of the treated bacterial spores were evaluated by using antibiotics (penicillin G, rifampicin and chloramphenicol) supplemented trypticase soy broth. In the chloramphenicol supplemented TSB, although the lag phase of bacterial spores treated by HPP with AlEW was not prolonged as compared with the normal-TSB, the decrease in growth rate during logarithmic phase and increase in maximum growth amount were observed. This result could suggest due to a damage by combined treatment of HPP with AlEW such as the inhibition of protein synthesis. Furthermore, the combined treatment with AlEW increased the catalase activity by 1.45, 1.63 and 4.25 times at 30, 80 and 100 MPa, respectively, as compared with solo-HPP, therefore the combined treatment could cause high oxidative stress on bacterial spores.

Highly efficient protein expression of Plasmodium vivax surface antigen, Pvs25, by silkworm and its biochemical analysis.

Plasmodium vivax ookinete surface protein, Pvs25, is a candidate for a transmission-blocking vaccine (TBV) for malaria. Pvs25 has four EGF-like domains containing 22 cysteine residues forming 11 intramolecular disulfide bonds, a structural feature that makes its recombinant protein expression difficult. In this study, we report the high expression of recombinant Pvs25 as a soluble form in silkworm, Bombyx mori. The Pvs25 protein was purified from hemolymphs of larvae and pupae by affinity chromatography. In the Pvs25 expressed by silkworm, no isoforms with inappropriate disulfide bonds were found, requiring no further purification step, which is necessary in the case of Pichia pastoris-based expression systems. The Pvs25 from silkworm was confirmed to be molecularly uniform by sodium dodecyl sulfate gel electrophoresis and size-exclusion chromatography. To examine the immunogenicity, the Pvs25 from B. mori was administered to BALB/c mice subcutaneously with oil adjuvant. The Pvs25 produced by silkworm induced potent and robust immune responses, and the induced antisera correctly recognized P. vivax ookinetes in vitro, demonstrating the potency of Pvs25 from silkworm as a candidate for a malaria TBV. To the best of our knowledge, this is the first study to construct a system for mass-producing malaria TBV antigens using silkworm.

Effect of the Combination of High Hydrostatic Pressure and Alkaline Electrolyzed Water on the Reduction of Heat Resistance of Bacterial Spores.

The effect of combined use of alkaline electrolyzed water （AlEW） on the reduction of heat resistance of bacterial spores by high hydrostatic pressure processing（ HPP） was investigated in this study. No reduction of heat resistance of bacterial spores, which was defined as the spore survival by heat treatment at 80℃ for 15 min, was observed by the treatment of single HPP with 30MPa at 50℃ even for 6 hours. However, a 3-log decrease in the viable bacterial spores was obtained by the combination of AlEW pretreatment with 1 hour of HPP treatment. An additional 2 hours duration of HPP treatment could inactivate more 2 logs of the viable bacterial spores. The obtained D value of bacterial spores treated by HPP was decreased to one-eighth by the pretreatment with AlEW when compared with the control sample. In case of the temperature during HPP treatment was 70℃, bacterial spores did not reduce its heat resistance with lower pressuring levels. In case of the temperature during HPP treatment is high with lower pressure levels, bacterial spores did not reduce its heat resistance even when AlEW was combined as the pretreatment. It was considered that the decrease in heat resistance by AlEW was resulted from the weakening of surface layer of spores by protein dissolution with alkaline substance. No clear effect of high negative redox potential, which is a unique property of AlEW, on the reduction of heat resistance was recognized.

Biomechanical study of the effect of degree of static compression of the spinal cord in ossification of the posterior longitudinal ligament.

OBJECT: The authors evaluated the biomechanical effect of 3 different degrees of static compression in a model of the spinal cord in order to investigate the effect of cord compression in patients with ossification of the posterior longitudinal ligament (OPLL). METHODS: A 3D finite element spinal cord model consisting of gray matter, white matter, and pia mater was established. As a simulation of OPLL-induced compression, a rigid plate compressed the anterior surface of the cord. The degrees of compression were 10, 20, and 40% of the anteroposterior (AP) diameter of the cord. The cord was supported from behind by the rigid body along its the posterior border, simulating the lamina. Stress distributions inside of the cord were evaluated. RESULTS: The stresses on the cord were very low under 10% compression. At 20% compression, the stresses on the cord increased very slightly. At 40% compression, the stresses on the cord became much higher than with 20% compression, and high stress distributions were observed in gray matter and the lateral and posterior funiculus. The stresses on the compressed layers were much higher than those on the uncompressed layer. CONCLUSIONS: The stress distributions at 10 and 20% compression of the AP diameter of the spinal cord were very low. The stress distribution at 40% compression was much higher. The authors conclude that a critical point may exist between 20 and 40% compression of the AP diameter of the cord such that when the degree of the compression exceeds this point, the stress distribution becomes much higher, and that this may contribute to myelopathy.

Biomechanical study of cervical flexion myelopathy using a three-dimensional finite element method.

OBJECT: The goal of this study was to perform a biomechanical study of cervical flexion myelopathy (CFM) using a finite element method. METHODS: A 3D finite element model of the spinal cord was established consisting of gray matter, white matter, and pia mater. After the application of semi-static compression, the model underwent anterior flexion to simulate CFM. The flexion angles used were 5 degrees and 10 degrees , and stress distributions inside the spinal cord were then evaluated. RESULTS: Stresses on the spinal cord were very low under semi-static compression but increased after 5 degrees of flexion was applied. Stresses were concentrated in the gray matter, especially the anterior and posterior horns. The stresses became much higher after application of 10 degrees of flexion and were observed in the gray matter, posterior funiculus, and a portion of the lateral funiculus. CONCLUSIONS: The 5 degrees model was considered to represent the mild type of CFM. This type corresponds to the cases described in the original report by Hirayama and colleagues. The main symptom of this type of CFM is muscle atrophy and weakness caused by the lesion of the anterior horn. The 10 degrees model was considered to represent a severe type of CFM and was associated with lesions in the posterior fand lateral funiculi. This type of CFM corresponds to the more recently reported clinical cases with combined long tract signs and sensory disturbance.

Effect of the wavelength of infrared heaters on the inactivation of bacterial spores at various water activities.

Bacterial spores (Bacillus subtilis subsp. subtilis NBRC 16183) inoculated onto a stainless steel Petri dish and treated at nine levels of water activity (a(w)) for 2 days were inactivated by infrared radiation heating (IRH) using three kinds of infrared heaters with different radiation spectra. The peak wavelengths used were 950, 1,100 and 1,150 nm. In general, the inactivating efficacy of IRH treatment against bacterial spores with shorter wavelength heater (950 nm) was greater than that with other heaters. The decimal reduction times (D value) calculated using the linear portion of survival curves were affected by both the initial a(w) values and the spectra of the infrared rays. Spores at approximately 0.9, 0.7 and 0.6 a(w) were most resistant to IRH at wavelengths of 950, 1,100 and 1,150 nm, respectively. The a(w) values that led to maximum D values for bacterial spores increased as the wavelength was shortened. Optimum a(w) values were identified for the inactivation of bacterial spores by IRH. Spore resistance to IRH could also be affected by the spectral characteristics of the infrared absorption, which varied with the a(w) of bacterial spores.