To analyse the metallo-β-lactamase (MBL) genes and their genetic environments in clinical isolates of Achromobacter xylosoxidans.
From January 2004 to December 2010, four MBL-producing, ...multidrug-resistant (MDR) A. xylosoxidans strains were isolated and analysed at a tertiary care university hospital in Japan. Species-level identification was confirmed by 16S ribosomal RNA gene sequencing. Molecular typing was performed using PFGE, the presence of MBL genes was detected using PCR, and integron gene cassettes were examined by cloning and sequence analysis of integron PCR products. The plasmids obtained from individual isolates were analysed based on EcoRI restriction patterns, Southern hybridizations using digoxigenin-labelled probes for bla(IMP-1) and bla(IMP-19) as well as conjugation and transformation experiments.
No clonal relationship was found among the four A. xylosoxidans isolates. Three isolates harboured bla(IMP-1) and one isolate harboured bla(IMP-19). These MBL genes were carried on class 1 integrons. Four different class 1 integron gene cassette arrays were found, including orf1-bla(IMP-1)-aacA4, bla(IMP-1)-bla(IMP-1)-nit1/nit2-catB3-bla(IMP-1)-bla(IMP-1), aacA4-bla(IMP-1) and bla(IMP-19). The restriction pattern of the plasmid DNA obtained from each isolate was unique and the hybridization analyses revealed the presence of each MBL gene within a plasmid. Moreover, all of the plasmids carrying an MBL gene could be transformed into Escherichia coli HST08.
This study provides genetic evidence for the existence of IMP-type MBL genes in MDR A. xylosoxidans isolates. Moreover, the present findings provide evidence that A. xylosoxidans can receive IMP-type MBL genes via plasmid-mediated transfer, which contributes to their carbapenem resistance.
The effects of microstructural characteristics (secondary dendrite arm spacing, SDAS) and Si- and Fe-based eutectic structures on the mechanical properties and failure behavior of an Al-Si-Cu alloy ...are investigated. Cast Al alloy samples are produced using a special continuous-casting technique with which it is easy to control both the sizes of microstructures and the direction of crystal orientation. Dendrite cells appear to grow in the casting direction. There are linear correlations between SDAS and tensile properties (ultimate tensile strength
σ
UTS
, 0.2 pct proof strength
σ
0.2
, and fracture strain
ε
f
). These linear correlations, however, break down, especially for
σ
UTS
vs
SDAS and
ε
f
vs
SDAS, as the eutectic structures become more than 3
μ
m in diameter, when the strength and ductility (
σ
UTS
and
ε
f
) decrease significantly. For eutectic structures larger than 3
μ
m, failure is dominated by the brittle eutectic phases, for which SDAS is no longer strongly correlated with
σ
UTS
and
ε
f
. In contrast, a linear correlation is obtained between
σ
0.2
and SDAS, even for eutectic structures larger than 3
μ
m, and the eutectic structure does not have a strong effect on yield behavior. This is because failure in the eutectic phases occurs just before final fracture.
In situ
failure observation during tensile testing is performed using microstructural and lattice characteristics. From the experimental results obtained, models of failure during tensile loading are proposed.
Purpose
The HyperEye Medical System (HEMS) uses indocyanine green (ICG) to visualize blood vessels in coronary artery bypass grafting (CABG). We performed quantitative HEMS assessment to detect ...grafts at risk of occlusion.
Methods
We assessed the HEMS angiograms of 177 grafts from 69 patients who underwent CABG and compared the results with those of fluoroscopic coronary angiography, by measuring the increasing rate of ICG intensity, average acceleration value, and time to peak luminance intensity.
Results
Grafts in the patent and failed groups showed significant differences in their increasing rate of intensity and average acceleration value. The average accelerations value of ICG intensity of internal thoracic artery (ITA) and saphenous vein (SV) grafts were 112.3 and 144.9 intensity/s
2
in the patent group, and 71.0 and 91.8 intensity/s
2
in the failed group. The time to peak luminance intensity was 1.7 and 1.4 s in the patent group and 2.3 and 1.9 s in the failed group; these values were not significantly different.
Conclusion
Significant reductions in the ICG intensity rate and average acceleration value can occur in failed grafts. Therefore, quantifiable changes in ICG intensity may help detect minute changes in blood flow.
When we teach motions to conventional industrial robot arms by Hand-Guiding instruction, the bad manipulability of the robot arms sometimes prevents us from teaching motions to them. In this paper, ...we apply the robot dual shell to an industrial robot arm. The robot dual shell has no contact with the robot arm. And the robot arm tracks the motion of the robot dual shell. Therefore introducing the robot dual shell improves the manipulability of the robot arm and maintains the range of movement of the robot arm. As a result, we can teach motions to the robot arm by Hand-Guiding instruction. Through the quantitative evaluation of manipulability and the practical operation, we show that the robot dual shell structure is indeed effective in improving manipulability.
A micro-focused X-ray beam with size ranging from 1 × 1 to 10 × 10 μm has been achieved at beamline BL32XU at SPring-8, Japan. Combining the available micro-beam with newly developed techniques has ...enabled efficient protein micro-crystallography.
Carbapenemase-producing Gram-negative bacilli have been a global concern over the past 2 decades because these organisms can cause severe infections with high mortality rates. Carbapenemase genes are ...often carried by mobile genetic elements, and resistance plasmids can be transferred through conjugation. We conducted whole-genome sequencing (WGS) to demonstrate that the same plasmid harboring a metallo-β-lactamase gene was detected in two different species isolated from a single patient. Metallo-β-lactamase-producing Achromobacter xylosoxidans (KUN4507), non-metallo-β-lactamase-producing Klebsiella pneumoniae (KUN4843), and metallo-β-lactamase-producing K. pneumoniae (KUN5033) were sequentially isolated from a single patient and then analyzed in this study. Antimicrobial susceptibility testing, molecular typing (pulsed-field gel electrophoresis and multilocus sequence typing), and conjugation analyses were performed by conventional methods. Phylogenetic and molecular clock analysis of K. pneumoniae isolates were performed with WGS, and the nucleotide sequences of plasmids detected from these isolates were determined using WGS. Conventional molecular typing revealed that KUN4843 and KUN5033 were identical, whereas the phylogenetic tree analysis revealed a slight difference. These two isolates were separated from the most recent common ancestor 0.74 years before they were isolated. The same resistance plasmid harboring blaIMP-19 was detected in metallo-β-lactamase-producing A. xylosoxidans and K. pneumoniae Although this plasmid was not self-transferable, the conjugation of this plasmid from A. xylosoxidans to non-metallo-β-lactamase-producing K. pneumoniae was successfully performed. The susceptibility patterns for metallo-β-lactamase-producing K. pneumoniae and the transconjugant were similar. These findings supported the possibility of the horizontal transfer of plasmid-borne blaIMP-19 from A. xylosoxidans to K. pneumoniae in a single patient.
Acute graft-versus-host disease (aGVHD) is a serious complication of allogeneic hematopoietic stem cell transplantation (HSCT). CC chemokine ligand 8 (CCL8) has been reported to be a potential ...molecular candidate for the diagnosis of aGVHD in both human and mouse systems. To elucidate a possible role of CCL8 in aGVHD pathology, we performed a murine allogeneic HSCT using CCL8 knockout (CCL8-/-) mice as recipients. When fully allogeneic marrow graft with splenocytes was transplanted, the early mortality and morbidity of the CCL8-/- host appeared to be markedly reduced compared to those of wild type control. This observation suggests that CCL8 is involved in the pathology of aGVHD and can be a potential biomarker as well as a therapeutic target for aGVHD.
Introduction: Although HSCT has been widely employed as a curative treatment for both malignant and nonmalignant diseases, a diagnostic test and therapy for GVHD have yet to be established. Control of aGVHD is a key process in order for HSCT to succeed. Consequently, there has been considerable effort to establish a simple noninvasive means of early and more precise diagnosis of aGVHD. We have previously shown that the levels of CCL8 are elevated in the plasma of murine aGVHD models and serum of humans with aGVHD (Hori T, et al. Blood. 2008;111:4403-4412), and closely correlate with the severity of aGVHD in mice (Yamamoto M, et al. Exp Hematol. 2011;39:1101-1112). To further examine the relationship between CCL8 and aGVHD, we tested a murine aGVHD model using CCL8-/- mice.
Materials and Methods: CCL8-/- mice were developed in C57BL/6 (B6) inbred mice through the zinc-finger nuclease system. Detailed characteristics of CCL8-/- mice are shown in the accompanying abstract in the current conference. Wild type B6 (control) or CCL8-/- mice were irradiated at 10 Gy and transferred with fully allogeneic transplants containing 2.0x107 bone marrow cells and 1.0x107 spleen cells of BALB/c. Kaplan-Meier survival curves were generated and analyzed by means of the log-rank test. Animal protocols were approved by the Ethics Review Committee for Animal Experimentation of Sapporo Medical University (approval number: 13-043).
Results: As shown in the figure, the early mortality of CCL8-/- recipients was obviously reduced compared to that of wild type B6 recipients. At day 10 after transplantation, only 33% of the wild type B6 mice survived (see rhombus in the figure), whereas 93% of the CCL8-/- mice survived (rectangle). CCL8-/- mice maintained this survival rate until day 28. Median survival time of wild type B6 and CCL8-/- mice was 9 days and 45 days, respectively. The difference was statistically significant (p <0.001, log-rank test).
Discussion: CCL8 deficiency in hosts obviously decreases an early mortality and greatly extends survival time in murine aGVHD. In addition, clinical symptoms and pathological findings of CCL8-/- recipients were dramatically improved at day 7 in comparison with those of wild type B6 recipients, showing that CCL8 plays a role in aGVHD morbidity. Taken together, these observations reflect that CCL8 is involved in the pathology of aGVHD.
The development of aGVHD can be classified into three phases: (1) host antigen presenting cell (APC) activation, (2) donor T cell activation, and (3) target tissue destruction (Ferrara JL, et al. Lancet. 2009;373:1550-1561). Plasma levels of interferon-gamma in CCL8-/- mice at day 5 of allogeneic HSCT were similar to those in wild type B6 mice, suggesting that donor T cells were activated in response to alloantigen presented by host APCs of CCL8-/- mice. The histopathological examination showed tissue injuries in target organs of aGVHD (skin, liver, gastrointestinal tract), but these were clearly attenuated compared with those of wild type B6 recipients. These results suggest that in CCL8-/- hosts the first and second phases of aGVHD occur similar to those in wild type B6 mice and tissue destruction in the third phase may be affected.
Collectively, these findings demonstrate that CCL8 has a strong correlation with the mortality and morbidity of aGVHD and can be a potential biomarker as well as a therapeutic target for aGVHD.
Display omitted
No relevant conflicts of interest to declare.
In Japan, X-ray structure analyses of molecules and particles from biology started in the 1970s. The structure analysis methods have been developed through the innovation of various techniques in ...advance, and have contributed for understanding the elementary and microscopic processes in life. Here we summarize briefly the history of X-ray structure analyses for structural biology in Japan and think about the prospect.
Abstract
Apolipoprotein E receptor 2 (Apo
ER
2) is a close homologue of low‐density lipoprotein receptor (
LDLR
) that mediates the endocytosis of ligands, including
LDL
particles.
LDLR
family ...members have been presumed to explore a large conformational space to capture ligands in the extended conformation at the cell surface. Ligands are subsequently released through a
pH
‐titrated structural transition to a self‐docked, contracted‐closed conformation. In addition to lipoprotein uptake, Apo
ER
2 is implicated in signal transduction during brain development through capture of the extracellular protein reelin. From crystallographic analysis, we determine that the full‐length Apo
ER
2 ectodomain adopts an intermediate contracted‐open conformation when complexed with the signaling‐competent reelin fragment, and we identify a previously unappreciated auxiliary low‐affinity binding interface. Based on mutational analyses, we propose that the
pH
shift during endocytosis weakens the affinity of the auxiliary interface and destabilizes the ligand–receptor complex. Furthermore, this study elucidates that the contracted‐open conformation of ligand‐bound Apo
ER
2 at neutral
pH
resembles the contracted‐closed conformation of ligand‐unbound
LDLR
at acidic
pH
in a manner suggestive of being primed for ligand release even prior to internalization.
Synopsis
image
Apo
ER
2 is a close homologue of the
LDL
receptor. The crystal structure of the Apo
ER
2 ectodomain in complex with the signaling‐competent fragment of reelin serves as a representative model for the
pH
‐dependent ligand uptake mechanism conserved within the
LDLR
family.
The structure of the ApoER2 ectodomain in complex with its physiological ligand has been determined by X‐ray crystallography.
The LA2 module of ApoER2 serves as a low‐affinity auxiliary interface for reelin and is expected to modulate ligand‐receptor interactions during endocytosis.
The conformation of ligand‐bound ApoER2 resembles that of the closed state of LDLR at acidic pH rather than that of the extended state at neutral pH, indicating that ApoER2 is primed for ligand release even before internalization.
Using brush tire model, it was proved that slip dissipation power on the tire contact patch at a tire slip angle is equal to the power loss due to the cornering resistance regardless of the ...distribution of adhesion and sliding regions. The analysis result also shows the dissipation power due to a camber angle is consistent with the cornering resistance obtained from the coordinate transformation. Therefore, the respective dissipation power can be calculated directly as a product of the generated cornering resistance and velocity. Finally, this paper classified the cornering resistance and dissipation power due to longitudinal slip ratio, lateral slip angle, and camber angle, respectively.
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