Development of Rapid and Specific Detection for the Human Aichivirus A Using the Loop-Mediated Isothermal Amplification from Water Samples.

Human Aichivirus A (AiV-A) is classified as a Kobuvirus, group IV positive sense single strand RNA viruses. The first outbreak of AiV-A was reported from Aichi Prefecture, Japan in 1989. AiV-A exists not only among clinical patients, such as diarrhea, but also in a variety of water environments, as its occurrence is reported across a wide geographical range, from developing to advanced countries. For diagnose of AiV-A from water samples, mostly polymerase chain reaction (PCR) system have been developed. However, loop-mediated isothermal amplification (LAMP) assay has not been applied. In this study, developed a LAMP method to achieve a rapid, specific and highly sensitive detection of AiV-A. The method developed in this study is aimed specifically at AiV-A. Through a specific and non-specific selection and sensitivity test process for the five prepared LAMP primer sets, one primer set and optimum reaction temperature were selected. A newly developed method was more rapid (approximately 2-8 h), specific and equivalent detection of AiV-A than with the conventional PCRs. In addition, confirm system of positive LAMP reaction was developed by using the restriction enzyme Aci I and Hae III. For evaluation and verification of developing LAMP assay, a was applied to twenty cDNA from groundwater samples. This study proved rapid and specific diagnosis of AiV-A from water samples, and it is also demanded to be applicable to other environmental, clinical and food samples.

Development and Optimization of a Reverse Transcription Hemi-Nested PCR Primer for the Detection of Potato Mop-Top Virus at Quarantine Inspection Sites in Korea.

Potato mop-top virus (PMTV) is classified as a plant quarantine virus in Korea. It is tested at import inspection sites. In this study, two sets of reverse transcription-hemi-nested polymerase chain reaction primers were developed to diagnose PMTV. In addition, modified- positive control plasmid was developed for the identification of false positive results in plant quarantine. The assay is expected to be useful for improving the detection and diagnosis of PMTV.

Development of a Specific Diagnostic System for Detecting Turnip Yellow Mosaic Virus from Chinese Cabbage in Korea.

Turnip yellow mosaic virus (TYMV) is a plant pathogenic virus transmitted mainly through its host Brassica spp. TYMV is originated from Europe. Its infection cases have been reported in Australia, Brazil, Turkey, and Japan. Symptoms similar to those of TYMV infections were also reported in Korea in 2012. In this study, we developed RT-PCR primer pairs that were highly sensitive for detecting TYMV. The developed RT-PCR primer pairs offered about 10-100 times stronger detection sensitivity compared to primer pairs previously used in Korea. As a result, a 491 bp TYMV-specific band was identified. The specific band was confirmed to be TYMV based on sequencing results and phylogenetic analysis. The RT-PCR primer pairs developed in this study can be used to rapidly and precisely diagnose TYMV in agricultural products such as Chinese cabbage and other crops infected by TYMV.

Development of PCR diagnostic system for detection of the seed-transmitted tobacco ringspot virus in quarantine.

Tobacco ringspot virus (TRSV) is a plant quarantine virus in Korea. As such, a TRSV examination is conducted when importing various crops. In this study, RT-PCR and nested PCR systems for TRSV detection in quarantine sites, and the modified-positive control plasmid for proving laboratory contamination and false positive reactions were developed. The developed diagnostic system was used to detect TRSV in the quarantine site. It revealed that from 2012 to August 2014, a total of 12 cases were detected in imported various crops. The system is expected to continue contributing to TRSV detection in plant quarantine.

Fat-forming variant of solitary fibrous tumour of the pleura: CT findings.

The fat-forming variant of solitary fibrous tumour (SFT) was previously called lipomatous haemangiopericytoma and is a rare variant of solitary fibrous tumour. It predominantly occurs in the deep soft tissues of the retroperitoneum and thigh. Only a handful of cases involving the perineum, spine, thoracic wall and pelvic cavity have been reported in the radiological literature and the fat-forming variant of SFT involving the pleura has not been previously reported. Herein, we report the CT findings of a case of the fat-forming variant of SFT involving the pleura that was treated by excision. Chest CT showed a large lobulated heterogeneous fatty mass with a multifocal enhancing soft-tissue component in the left lower hemithorax. Although rare, the fat-forming variant of SFT of the pleura should be added to the differential diagnosis of fat-containing pleural soft-tissue tumours.

A case of a giant mucocoele of the appendiceal stump presented with a palpable mass in the right thigh: pre-operative diagnosis based on characteristic multidetector CT findings.

The pre-operative diagnosis of a mucocoele of the appendiceal stump (MAS) may be difficult owing to rarity and non-specific clinical presentation. However, a pre-operative diagnosis of a MAS is important to prevent widespread dissemination by inadvertent spillage of mucous contents. We describe a case of a MAS presenting with a palpable mass in the right thigh in which a pre-operative diagnosis was made by characteristic multidetector CT (MDCT) findings.

The role of hand-assisted laparoscopic distal gastrectomy for distal gastric cancer.

BACKGROUND: The aim of this study was to evaluate the role of hand-assisted distal gastrectomy (HALDG) for gastric cancer. METHODS: This study prospectively enrolled 16 patients who underwent HALDG for early gastric cancer and matched them individually by sex, age, and body mass index to patients who underwent laparoscopically assisted distal gastrectomy (LADG) or open distal gastrectomy (ODG). Surgical outcomes were compared among the surgical methods. RESULTS: The mean operating time was the longest for the HALDG group, whereas wound size of the HALDG group was intermediate between that of the LADG and the ODG groups. The other surgical outcomes, such as the number of harvested lymph nodes, were not different among the groups. CONCLUSIONS: According to the findings, HALDG may not be as beneficial for patients with early gastric cancer as has been previously suggested. However, because of easier hand-eye coordination, HALDG may be an excellent bridge learning technique as a surgeon gains experience in laparoscopic gastrectomy.

Bone intrinsic material properties in three inbred mouse strains.

This study assessed genetically based differences in intrinsic material properties of both cortical and cancellous bone in adult females of three inbred mouse strains [C57BL/6J (B6), DBA/2J (D2), C3H/HeJ (C3)]. These mouse strains have previously been shown to differ in bone mineral content (BMC) and density (BMD). Distal femoral cancellous bone and midshaft cortical bone in femurs and tibias were assessed for intrinsic material properties using nanoindentation technique. The intrinsic material properties tested were modulus (E(b)) and hardness (H) of the midshaft femoral and tibial cortical bone cross sections and of cancellous bone in the distal femur. Both femoral and tibial cortical bone intrinsic material properties were different among the three inbred mouse strains. Femoral modulus and tibial hardness in cortical bone and hardness in cancellous bone were either greatest or showed greater trends in C3 mice as compared to both D2 and B6. Cancellous bone modulus was similar among the three mouse strains. With the exception of the D2 mice, the femoral and tibial cortical modulus were similar within each mouse strain. The tibial cortical modulus was smaller than the femoral cortical modulus for D2 mouse strain. The cortical hardness was greater in tibiae compared with that in femora within each mouse strain. The nanoindentation data suggest that cortical and cancellous intrinsic material properties are influenced by the genetic background of the inbred mouse strains. The inbred mouse strain-related intrinsic material property phenotype can be used to locate responsible quantitative trait loci (QTLs) in future studies of recombinant inbred mouse strains.

Atomic force microscopy and nanoindentation characterization of human lamellar bone prepared by microtome sectioning and mechanical polishing technique.

Surface topography, microstructure, and micromechanical properties of human lamellar bone were characterized by atomic force microscopy and nanoindentation. The lamellar bone surfaces were prepared by two different methods: microtome sectioning and mechanical polishing. The lamellar bone surfaces prepared by mechanical polishing revealed that thin lamellae formed depressions approximately 200 nm deep, whereas the surfaces prepared by microtome sectioning were flat. Atomic force microscopy surface topographic images at higher magnification showed differences between thick and thin lamellae in polished samples, but these differences were less pronounced in microtomed samples. Roughness measurements confirmed that there was a significant difference between thick (21.0 nm) and thin lamellae (8.3 nm) in polished samples (p < 0.001). The difference in surface roughness between thick (13.9 nm) and thin lamellae (12.7 nm) in microtomed sample was statistically insignificant (p = 0.74). Higher elastic modulus values were observed for thick lamella in microtomed samples compared with that of thin lamellae, whereas measured elastic modulus differences between thick and thin lamellae in polished samples were found to be statistically insignificant.

Congenital lack of COX-2 affects mechanical and geometric properties of bone in mice.

We compared the mechanical properties of bones from mice lacking either a functional cycloxygenase-1 (C57BL6/DBA COX-1-/-; n = 9) or COX-2 (C57BL6/DBA COX-2-/-; n = 9) gene and wild type mice (C57BL6/DBA; n = 10). Twenty-eight right femora from 3-month-old male mice were used to determine bulk structural and material properties of bone by three-point bending. Bone matrix properties were also measured by nanoindentation to access the changes in bulk mechanical properties due to changes in bone matrix or bone geometry. The bulk material properties (elastic modulus, P < 0.05; ultimate stress, P < 0.01) of COX-2-/- bones were lower than those of wild-type mice whereas the bulk structural properties (stiffness, P > 0.2; breaking force, P > 0.1) were similar to those of the wild-type mice. COX-2-/- mice had a longer moment of inertia but their cortical bones were thinner and contained many more intra-cortical pores compared with the bones of the other two groups. Finally, the bone matrix properties of COX-1-/- mice, COX-2-/- mice and their heterozygous littermates were similar to those of C57BL6/DBA wild-type mice.

Anisotropic properties of human tibial cortical bone as measured by nanoindentation.

The purpose of this study was to investigate the effects of elastic anisotropy on nanoindentation measurements in human tibial cortical bone. Nanoindentation was conducted in 12 different directions in three principal planes for both osteonic and interstitial lamellae. The experimental indentation modulus was found to vary with indentation direction and showed obvious anisotropy (one-way analysis of variance test, P < 0.0001). Because experimental indentation modulus in a specific direction is determined by all of the elastic constants of cortical bone, a complex theoretical model is required to analyze the experimental results. A recently developed analysis of indentation for the properties of anisotropic materials was used to quantitatively predict indentation modulus by using the stiffness matrix of human tibial cortical bone, which was obtained from previous ultrasound studies. After allowing for the effects of specimen preparation (dehydrated specimens in nanoindentation tests vs. moist specimens in ultrasound tests) and the structural properties of bone (different microcomponents with different mechanical properties), there were no statistically significant differences between the corrected experimental indentation modulus (Mexp) values and corresponding predicted indentation modulus (Mpre) values (two-tailed unpaired t-test, P > 0.5). The variation of Mpre values was found to exhibit the same trends as the corrected Mexp data. These results show that the effects of anisotropy on nanoindentation measurements can be quantitatively evaluated.

Effect of collagen and mineral content on the high-frequency ultrasonic properties of human cancellous bone.

The technology surrounding ultrasonic bone assessment is evolving rapidly as investigators explore the utility of new ultrasonic parameters and different ultrasonic frequencies. This study had three main goals. The first was to perform in vitro measurements of the speed of sound (SOS) and normalized broadband ultrasonic attenuation (nBUA) in specimens of normal human cancellous bone using a 2.25 MHz broadband measurement system. The second was to explore the utility of a backscatter-based parameter called apparent integrated backscatter (AIB). The third goal was to investigate the roles that collagen and mineral content play in affecting each of these three ultrasonic parameters. This was accomplished by chemically treating the specimens to remove one or the other of these two important constituents of bone. Our results showed that in most cases SOS and nBUA correlated well (p < 0.05) with bone mineral density (BMD) as measured by quantitative computed tomography (QCT). In contrast, AIB did not correlate strongly with BMD. When the specimens were demineralized, decreases were produced in SOS (19-39%) and nBUA (44-58%). Changes produced in AIB were not significant except along the superoinferior direction, in which a 12% decrease was measured. When the specimens were decollagenized, decreases were produced in SOS (10-12%). In contrast, increases were produced in both nBUA (35-77%) and AIB (14-15%). From this study we conclude that high-frequency ultrasonic measurements may yield useful information about the content and organization of both collagen and mineral in cancellous bone.

Microstructural elasticity and regional heterogeneity in human femoral bone of various ages examined by nano-indentation.

The elastic modulus and hardness of secondary osteonal and interstitial bone was examined through the thickness of the cortex of human femora of various ages by nano-indentation. There was a clear difference between the stiffness and hardness of secondary osteonal and interstitial bone, the latter being stiffer (F(1,48)=56.0, P<0.001). There were some differences between the bones of different subjects; however, there were no differences that could be reliably associated with the chronological age of the subject, or with differences in location through the thickness of the cortex (F(2,48)=0.21, P=0.810). Previous studies have been equivocal in relating changes in the macroscopic 'composite' material stiffness of bone to the age of the individual. By combining the results of the nano-tests with histological measures, we were able to produce a good relationship of the microstructural properties at the matrix level with the bending modulus of whole bone (R(2)=0.88, P<0.001) and this improved further by taking into account the age of the individual (R(2)=0.94, P<0.001). Our results suggest that using differences in the volumetric proportions of secondary osteons versus interstitial bone, and the properties of these elements/structures in isolation may be a more accurate method of determining differences in elastic modulus of whole bone between individuals of various ages.

Carcinoma originating from aberrant breast tissue of the right upper anterior chest wall : a case report.

Aberrant breast tissue is usually found in proximity to the normal breast, that is, in the axillary, sternal or clavicular regions. Carcinoma occurs more frequently in the aberrant tissue of the axilla than the extra-axillary site though the overall incidence of tumors of aberrant breast tissue is low. To our knowledge, studies regarding the carcinoma of aberrant breast tissue of the extra-axillary site have been reported rarely. Here we report a recent case of carcinoma originating from the extra-axillary aberrant breast tissue, presenting as a subcutaneous nodule on the right upper anterior chest wall. It is suggested that subcutaneous nodules of uncertain origin around the periphery of the breast should be suspected for breast carcinoma as a differential diagnosis and treated properly.

Effects of anisotropy on elastic moduli measured by nanoindentation in human tibial cortical bone.

Many biological materials are known to be anisotropic. In particular, microstructural components of biological materials may grow in a preferred direction, giving rise to anisotropy in the microstructure. Nanoindentation has been shown to be an effective technique for determining the mechanical properties of microstructures as small as a few microns. However, the effects of anisotropy on the properties measured by nanoindentation have not been fully addressed. This study presents a method to account for the effects of anisotropy on elastic properties measured by nanoindentation. This method is used to correlate elastic properties determined from earlier nanoindentation experiments and from earlier ultrasonic velocity measurements in human tibial cortical bone. Also presented is a procedure to determine anisotropic elastic moduli from indentation measurements in multiple directions.

Correlations between osteocalcin content, degree of mineralization, and mechanical properties of C. carpio rib bone.

Osteocalcin is one of the most abundant noncollagenous proteins in bone. It is strongly associated with the mineral phase of bone, and has long been associated as a marker of bone turnover. However, its relationship to bone composition, strength, and structure is unclear. Carp rib bone is an excellent model for the study, because osteocalcin represents almost 60% of the total extractable noncollagenous proteins found in it. Because of the abundance of osteocalcin relative to other extractable proteins, any changes in the properties of carp rib bone would be more likely influenced by the osteocalcin concentration. To test the hypotheses that the concentration of osteocalcin is reflected in other properties of bone, the correlations between the osteocalcin concentration and the mineral content, microstructural properties, and physical characteristics of the bone mineral crystals were determined utilizing radioimmunoassay (RIA), spectrophotometry, nanoindentation, and small-angle X-ray scattering (SAXS) techniques, respectively. Osteocalcin concentration was found to be correlated to the molar Ca/P ratio and inversely correlated to the elastic modulus and hardness in the longitudinal plane. This study provides evidence for a putative relationship between the concentration of osteocalcin and the microstructural mechanical properties of bone. Correlations were also found between the mechanical properties in the longitudinal plane and both the phosphate content and the molar Ca/P ratio. However, no relationships could be identified between osteocalcin concentration and several parameters of bone crystals, as determined by SAXS.

Ultrasonic characterization of the curing process of hydroxyapatite-modified bone cement.

Ultrasonic parameters such as velocity of sound and broad-band ultrasonic attenuation (BUA) are sensitive to changes in the viscoelastic properties of a material. Bone cement undergoes changes is these properties as it cures. By monitoring the propagation of ultrasonic pulses through a sample of curing bone cement, the curing reaction of polymethylmethacrylate-based (PMMA) bone cement was investigated for hydroxyapatite (HA) concentrations of 0, 10, and 30% (by weight). As the material hardens, the velocity of sound increases by 70%. BUA shows a large peak at the midpoint of the velocity transition. These data are used to compare the cure time and cure duration for PMMA bone cement mixed with hydroxyapatite particles. Measurements of the final sound velocity and BUA were also performed to investigate the mechanical properties of the fully cured cement, and to compare to compression testing data. This is the first time the curing process of bone cement has been investigated as a function of hydroxyapatite concentration. Results indicate that the cure time is not significantly affected by the addition of HA particles, and that both velocity of sound and BUA are sensitive to the curing process.

The anisotropic Young's modulus of equine secondary osteones and interstitial bone determined by nanoindentation.

The equine radius is a useful subject for examining the adaptation of bone histology to loading because in life the anterior cortex is loaded almost entirely in tension, the posterior cortex in compression. The histology of the two cortices is correspondingly different, the osteones and the interstitial lamellae in the posterior cortex having a more transversely oriented fibre arrangement than those in the anterior cortex. Presumably as a result of this histological difference, the posterior cortex is stronger in compression than the anterior cortex; the anterior cortex is stronger in tension than the posterior cortex. We here use nanoindentation to examine how the Young's modulus of elasticity of secondary osteones and interstitial lamellae in the anterior and posterior cortices varied as a function of angle. The anterior osteones were stiffer than the posterior osteones when tested in the direction parallel to the bone's long axis, but became progressively relatively less stiff as the angle increased; at 90 degrees, they were less stiff than the posterior osteones. Although the interstitial lamellae were stiffer than their neighbouring osteones, the same relationship between anterior and posterior interstitial lamellae as a function of angle was found as for the osteones. The anisotropy of these Young's moduli determined by nanoindentation shows a close relationship with what was to be expected from the histological findings.

A lead-absorbing protein with superoxide dismutase activity from Streptomyces subrutilus.

A lead binding protein was purified from the culture filtrate of Streptomyces subrutilus P5. The subunit and native molecular weights were estimated to be 28 and 55 kDa, respectively, indicating that the protein was composed of two identical subunits. The inhibition pattern, the metal content analysis and the EPR spectrum confirmed that the protein was a superoxide dismutase containing Fe and Zn (FeZnSOD). The protein precipitated immediately upon mixing with lead ions and the saturation number of lead ions was about 1100 lead atoms per subunit. Using this property, lead ions could be effectively removed from solutions.