Unraveling Antimicrobial Resistance Genes and Phenotype Patterns among Enterococcus faecalis Isolated from Retail Chicken Products in Japan

Multidrug-resistant enterococci are considered crucial drivers for the dissemination of antimicrobial resistance determinants within and beyond a genus. These organisms may pass numerous resistance determinants to other harmful pathogens, whose multiple resistances would cause adverse consequences. Therefore, an understanding of the coexistence epidemiology of resistance genes is critical, but such information remains limited. In this study, our first objective was to determine the prevalence of principal resistance phenotypes and genes among Enterococcus faecalis isolated from retail chicken domestic products collected throughout Japan. Subsequent analysis of these data by using an additive Bayesian network (ABN) model revealed the co-appearance patterns of resistance genes and identified the associations between resistance genes and phenotypes. The common phenotypes observed among E. faecalis isolated from the domestic products were the resistances to oxytetracycline (58.4%), dihydrostreptomycin (50.4%), and erythromycin (37.2%), and the gene tet(L) was detected in 46.0% of the isolates. The ABN model identified statistically significant associations between tet(L) and erm(B), tet(L) and ant(6)-Ia, ant(6)-Ia and aph(3’)-IIIa, and aph(3’)-IIIa and erm(B), which indicated that a multiple-resistance profile of tetracycline, erythromycin, streptomycin, and kanamycin is systematic rather than random. Conversely, the presence of tet(O) was only negatively associated with that of erm(B) and tet(M), which suggested that in the presence of tet(O), the aforementioned multiple resistance is unlikely to be observed. Such heterogeneity in linkages among genes that confer the same phenotypic resistance highlights the importance of incorporating genetic information when investigating the risk factors for the spread of resistance. The epidemiological factors that underlie the persistence of systematic multiple-resistance patterns warrant further investigations with appropriate adjustments for ecological and bacteriological factors.     

Synthesis of CDP-diacylglycerol by rat liver rough microsomes as visualized by electron microscopic autoradiography: relationship to GTP-stimulated membrane fusion

Using detergent-free conditions of incubation for the analysis of liponucleotide synthesis, we compared GTP-dependent formation of CDP-diacylglycerol (CDP-DG) and membrane fusion in RNA-depleted rough microsomes from rat liver. After incubation of stripped rough microsomes (SRM) in the presence of GTP and [5-3H]-CTP, radioactivity was recovered in lipid extracts and identified by thin-layer chromatography as a single spot which co-migrated with CDP-DG. The nucleotide requirement for CDP-DG synthesis and that for membrane fusion were observed to be identical. We next carried out an electron microscopic autoradiographic analysis on incubated membranes to determine the site of incorporation of [5-3H]-CTP. Silver grains were observed directly over the unilamellar membranes of natural vesicles. In confirmation of the biochemical data, quantitation of silver grain density indicated more grains over membranes incubated in the presence of GTP than over those incubated in the absence of this nucleotide. For membranes incubated in the presence of GTP, the grain density was similar over fused and unfused membranes in the same preparation. When SRM were incubated with the enzyme co-factors required for synthesis of phosphatidylinositol, a GTP-independent membrane fusion was observed by both transmission and freeze-fracture electron microscopy. Together with the biochemical and autoradiographic data, this suggests that phospholipid metabolism may be activated by GTP and lead to the fusion of RER membrane.     

CA9 transcriptional expression determines prognosis and tumour grade in tongue squamous cell carcinoma patients

CA9 is a member of the carbonic anhydrases’ family, that is often expressed in cancer cells under hypoxic condition. However, the role of CA9 in the molecular mechanisms of tongue squamous cell carcinoma (TSCC) pathogenesis remains unclear. CA9 expression was analysed using the TCGA database, and its influence on survival was performed using Kaplan-Meier, LASSO and COX regression analyses. The correlation between CA9 and immune infiltration was investigated by CIBERSORT and ESTIMATE. Moreover, the relationship between CA9 expression and downstream molecular regulation pathways was analysed by GSEA, GO and WGCNA. CA9 expression correlated with clinical prognosis and tumour grade in TSCC. Moreover, CA9 expression potentially contributes to the regulation of cancer cell differentiation and mediates tumour-associated genes and signalling pathways, including apoptosis, hypoxia, G2M checkpoint, PI3K/AKR/mTOR signalling and TGF-beta signalling pathways. However, the follicular helper T cells, regulatory T cells, immune and stromal scores showed no significance between high and low CA9 expression groups. These findings suggested that CA9 plays a critical role of TSCC prognosis and tumour grade. CA9 expression significantly correlated with the regulation of cell differentiation, various oncogenes and cancer-associated pathways.     

The dynamical structure of the RNA in alfalfa mosaic virus studied by 31P-nuclear magnetic resonance

The structure of the viral RNA in alfalfa mosaic virus (AlMV) was investigated by means of 31P-nuclear magnetic resonance (NMR). It was found that the 31P-NMR line width of AlMV Top a particles is significantly smaller than that of the larger Bottom particles. At low temperatures, the totational correlation time of the 31P nuclei essentially equals the tumbling rate of the virus particle, indicating that the RNA is contained rigidly inside the virion. At more elevated temperatures, the NMR line width sharpens more than expected on the basis of viscosity changes and the RNA exhibits internal mobility. The occurrence of internal mobility is paralleled by an increased internal mobility of the N-terminal part of the coat protein, as could be observed by 1H-NMR spectroscopy. The influence of EDTA on the 31P-NMR line width appeared to be negligible, which is in agreement with the idea that AlMV does not 'swell' like several other RNA-containing plant viruses.     

Understanding Performance Degradation in Cation‐Disordered Rock‐Salt Oxide Cathodes

The collective redox activities of transition‐metal (TM) cations and oxygen anions have been shown to increase charge storage capacity in both Li‐rich layered and cation‐disordered rock‐salt cathodes. Repeated cycling involving anionic redox is known to trigger TM migration and phase transformation in layered Li‐ and Mn‐rich (LMR) oxides, however, detailed mechanistic understanding on the recently discovered Li‐rich rock‐salt cathodes is largely missing. The present study systematically investigates the effect of oxygen redox on a Li_1.3Nb_0.3Mn_0.4O₂ cathode and demonstrates that performance deterioration is directly correlated to the extent of oxygen redox. It is shown that voltage fade and hysteresis begin only after initiating anionic redox at high voltages, which grows progressively with either deeper oxidation of oxygen at higher potential or extended cycling. In contrast to what is reported on layered LMR oxides, extensive TM reduction is observed but phase transition is not detected in the cycled oxide. A densification/degradation mechanism is proposed accordingly which elucidates how a unique combination of extensive chemical reduction of TM and reduced quality of the Li percolation network in cation‐disordered rock‐salts can lead to performance degradation in these newer cathodes with 3D Li migration pathways. Design strategies to achieve balanced capacity and stability are also discussed.     

Automated high-performance liquid chromatographic method for the determination of antipyrine and its metabolites in urine : Some preliminary results obtained from smokers and non-smokers

A simple, accurate and fully automated high-performance liquid-chromatographic method was developed for the simultaneous determination of antipyrine (AP), 3-hydroxymethylantipyrine (3HMA), 4-hydroxyantipyrine (4OHA) and norantipyrine (NORA) in urine. This method requires no extraction step and only one chromatographic run with the use of a reversed-phase system. The coefficient of variation (%) (n = 8 each) was: 4.14 for AP, 2.31 for 3HMA, 3.48 for 4OHA, and 2.71 for NORA. The method was applied to studies on AP metabolism in three smokers and three non-smokers who received an oral 10 mg/kg dose of AP. These preliminary results suggest that smokers appear to excrete more 4OHA and NORA in the urine than non-smokers.     

Naloxone reversal of ischemic arrhythmia is stereospecific and suggests role of endogenous opioid peptides in ischemic heart disease.

The effects of the stereoisomers of naloxone during myocardial ischemia were studied. (-)-Naloxone (but not the (+)-isomer naloxone) attenuated the ischemia-induced cardiac arrhythmias, hypotension, and bradycardia that result from coronary artery occlusion in anesthetized rats. From these findings, it may be inferred that endogenous opioid peptides may play a role in the pathophysiology of myocardial ischemia. It is also suggested that naloxone may have therapeutic value in the prevention and treatment of ischemic heart disease.     

Role of reversing factor in the inhibition of protein synthesis initiation by oxidized glutathione

The inhibitions of protein synthesis initiation in heme-deficient reticulocyte lysates and in GSSG-treated hemin-supplemented lysates are both characterized by the activation of heme-regulated eIF-2 alpha kinase, which phosphorylates the alpha-subunit of eukaryotic initiation factor (eIF-2). In both inhibitions, the accumulation of eIF phosphorylated in alpha-subunit (eIF-2(alpha P)) leads to the sequestration of reversing factor (RF) in a phosphorylated 15 S complex, RF.eIF-2(alpha P), in which RF is nonfunctional. A sensitive assay for the detection of endogenous RF activity in protein-synthesizing lysates indicates that, in GSSG-inhibited (1 mM GSSG) lysates, RF is more profoundly inhibited than in heme-deficient lysates. RF inactivation in GSSG-induced inhibition appears to be due to two separate but additive effects: (i) the formation of the phosphorylated 15 S RF complex, RF.eIF-2(alpha P), and (ii) the formation of disulfide complexes which inhibit RF activity. Both inhibitory effects are overcome by catalytic levels of exogenous RF which permits the resumption of protein synthesis. RF activity and protein synthesis in GSSG-inhibited lysates are efficiently restored by the delayed addition of glucose-6-P or 2-deoxyglucose-6-P (1 mM). The rescue of protein synthesis by hexose phosphate (1 mM) is proportional to the extent of RF recovery and is due in part to NADPH generation; even at levels of hexose phosphate (50 microM) too low to support protein synthesis, partial restoration of RF activity occurs due to increased NADPH/NADP+ ratios. The ability of dithiothreitol (1 mM) to restore RF activity in GSSG-treated but not heme-deficient lysates also provides evidence for a reducing mechanism which functions at the level of RF. The results suggest that NADPH plays a role in the maintenance of sulfhydryl groups essential for RF activity.     

A quantitative estimation of sequential foliage development and fertility inDryopteris crassirhizoma

The foliage development ofDryopteris crassirhizoma Nakai was quantitatively estimated by measurements of shape and size of leaves from different developmental stages of sporophytes, to lead to an understanding of the life history characteristics of the species. The number of midrib branches (NV, number of veins) of the leaf corresponds to the leaf-shape complexity (DI, dissection index; shape complexity from a circle) and length of leafblade (BL). Some quantitative characters, such as leaf uniformity (decrease in NV variation), changes in shape and increase in number of leaves, vary progressively during foliage formation. The sequence of foliage development can be quantified using the parameter NV: for example, 15-NV for leaf uniformity, 30-NV for leaf-shape change from triangular to oblanceolate, 60-NV for increase in leaf number and leaf fertility in the course of sporophyte ontogeny. Contribution No. 3297 from the Institute of Low Temperature Science, Hokkaido University.