生防细菌CNY-04筛选、鉴定及其对灰霉病菌的防效

【目的】为研究土壤细菌对蔬菜灰霉病的生防价值, 从辽宁、山东等地区的蔬菜种植基地采集土壤样本56份, 分离、筛选出对灰霉病具有稳定拮抗作用的细菌9株。【方法】采用平板对峙培养法进行初筛、复筛, 用抑菌圈法测定其抑菌效果, 并进行离体果实试验验证其对蔬菜灰霉病的防治效果, 通过形态学特征、生理生化特征及16S rRNA基因序列分析研究其分类地位。【结果】细菌CNY-04对蔬菜灰霉病的拮抗能力最强且遗传稳定, 抑菌圈直径达到34 mm; 初步鉴定该菌株为格氏沙雷菌(Serratia grimesii), 尚未见该菌在生防上的报道; CNY-04液体菌剂对离体番茄果实灰霉病的防效为69.23%, 50%多菌灵防效为75.39%, 24 h时接种CNY-04处理的番茄发病率为40.0%, 而48 h时接种处理的发病率为51.1%。【结论】CNY-04是一株较为理想的拮抗菌, 丰富了生防资源。     

Malaria parasite liver stages render host hepatocytes susceptible to mitochondria-initiated apoptosis

Intracellular eukaryotic parasites and their host cells constitute complex, coevolved cellular interaction systems that frequently cause disease. Among them, Plasmodium parasites cause a significant health burden in humans, killing up to one million people annually. To succeed in the mammalian host after transmission by mosquitoes, Plasmodium parasites must complete intracellular replication within hepatocytes and then release new infectious forms into the blood. Using Plasmodium yoelii rodent malaria parasites, we show that some liver stage (LS)-infected hepatocytes undergo apoptosis without external triggers, but the majority of infected cells do not, and can also resist Fas-mediated apoptosis. In contrast, apoptosis is dramatically increased in hepatocytes infected with attenuated parasites. Furthermore, we find that blocking total or mitochondria-initiated host cell apoptosis increases LS parasite burden in mice, suggesting that an anti-apoptotic host environment fosters parasite survival. Strikingly, although LS infection confers strong resistance to extrinsic host hepatocyte apoptosis, infected hepatocytes lose their ability to resist apoptosis when anti-apoptotic mitochondrial proteins are inhibited. This is demonstrated by our finding that B-cell lymphoma 2 family inhibitors preferentially induce apoptosis in LS-infected hepatocytes and significantly reduce LS parasite burden in mice. Thus, targeting critical points of susceptibility in the LS-infected host cell might provide new avenues for malaria prophylaxis.     

Intravascular heavy chain-modification of hyaluronan during endotoxic shock

During inflammation, the covalent linking of the ubiquitous extracellular polysaccharide hyaluronan (HA) with the heavy chains (HC) of the serum protein inter alpha inhibitor (IαI) is exclusively mediated by the enzyme tumor necrosis factor α (TNFα)-stimulated-gene-6 (TSG-6). While significant advances have been made regarding how HC-modified HA (HC-HA) is an important regulator of inflammation, it remains unclear why HC-HA plays a critical role in promoting survival in intraperitoneal lipopolysaccharide (LPS)-induced endotoxemia while exerting only a modest role in the outcomes following intratracheal exposure to LPS. To address this gap, the two models of intraperitoneal LPS-induced endotoxic shock and intratracheal LPS-induced acute lung injury were directly compared in TSG-6 knockout mice and littermate controls. HC-HA formation, endogenous TSG-6 activity, and inflammatory markers were assessed in plasma and lung tissue. TSG-6 knockout mice exhibited accelerated mortality during endotoxic shock. While both intraperitoneal and intratracheal LPS induced HC-HA formation in lung parenchyma, only systemically-induced endotoxemia increased plasma TSG-6 levels and intravascular HC-HA formation. Cultured human lung microvascular endothelial cells secreted TSG-6 in response to both TNFα and IL1β stimulation, indicating that, in addition to inflammatory cells, the endothelium may secrete TSG-6 into circulation during systemic inflammation. These data show for the first time that LPS-induced systemic inflammation is uniquely characterized by significant vascular induction of TSG-6 and HC-HA, which may contribute to improved outcomes of endotoxemia.     

Analysis of simulated NMR order parameters for lipid bilayer structure determination

The conventional formula for relating CD2 average order parameters to average methylenic travel is flawed when compared to molecular dynamics simulations of dipalmitoylphosphatidylcholine. Inspired by the simulated probability distribution functions, a new formula is derived that satisfactorily relates these quantities. This formula is used to obtain the average chain length , and the result agrees with the direct simulation result for . The simulation also yields a hydrocarbon thickness 2. The result = is consistent with a model of chain packing with both early chain termination and partial interdigitation of chains from opposing monolayers. The actual simulated area per lipid is easily obtained from the order parameters. However, when this method is applied to NMR order parameter data from dimyristoylphosphatidylcholine, the resulting is 10% larger than the currently accepted value.     

Interpretation of small angle X-ray measurements guided by molecular dynamics simulations of lipid bilayers

Reconstruction and interpretation of lipid bilayer structure from X-ray scattering often rely on assumptions regarding the molecular distributions across the bilayer. It is usually assumed that changes in head-head spacings across the bilayer, as measured from electron density profiles, equal the variations in hydrocarbon thicknesses. One can then determine the structure of a bilayer by comparison to the known structure of a lipid with the same headgroup. Here we examine this procedure using simulated electron density profiles for the benchmark lipids DMPC and DPPC. We compare simulation and experiment in both real and Fourier space to address two main aspects: (i) the measurement of head-head spacings from relative electron density profiles, and (ii) the determination of the absolute scale for these profiles. We find supporting evidence for the experimental procedure, thus explaining the robustness and consistency of experimental structural results derived from electron density profiles. However, we also expose potential pitfalls in the Fourier reconstruction that are due to the limited number of scattering peaks. Volumetric analysis of simulated bilayers allows us to propose an improved, yet simple method for scale determination. In this way we are able to remove some of the restrictions imposed by limited scattering data in constructing reliable electron density profiles.     

Heme oxygenase-1 inhibits TNF-alpha-induced apoptosis in cultured fibroblasts

Heme oxygenase (HO)-1 catalyzes the oxidative cleavage of heme to yield equimolar amounts of biliverdin, iron, and carbon monoxide. HO-1 is a stress response protein, the induction of which is associated with protection against oxidative stress. The mechanism(s) of protection is not completely elucidated, although it is suggested that one or more of the catalytic by-products provide antioxidant functions either directly or indirectly. The involvement of reactive oxygen species in apoptosis raised the question of a possible role for HO-1 in programmed cell death. Using the tetracycline-regulated expression system, we show here that conditional overexpression of HO-1 prevents tumor necrosis factor-alpha-induced apoptosis in murine L929 fibroblasts. Inhibition of apoptosis was not observed in the presence of tin protoporphyrin, a specific inhibitor of HO activity, and in cells overexpressing antisense HO-1. Interestingly, exogenous administration of a low concentration of carbon monoxide also prevented tumor necrosis factor-alpha-induced apoptosis in L929 fibroblasts. Inhibition of tumor necrosis factor-alpha-induced apoptosis by HO-1 overexpression was reversed by 1H-(1,2, 4)oxadiazolo(4,3-a)quinoxalin-1-one, an inhibitor of guanylate cyclase, which is a target enzyme for carbon monoxide. Taken together, our data suggest that the antiapoptotic effect of HO-1 may be mediated via carbon monoxide.     

Area per lipid and acyl length distributions in fluid phosphatidylcholines determined by (2)H NMR spectroscopy

Deuterium ((2)H) NMR spectroscopy provides detailed information regarding the structural fluctuations of lipid bilayers, including both the equilibrium properties and dynamics. Experimental (2)H NMR measurements for the homologous series of 1, 2-diacyl-sn-glycero-3-phosphocholines with perdeuterated saturated chains (from C12:0 to C18:0) have been performed on randomly oriented, fully hydrated multilamellar samples. For each lipid, the C-D bond order parameters have been calculated from de-Paked (2)H NMR spectra as a function of temperature. The experimental order parameters were analyzed using a mean-torque potential model for the acyl chain segment distributions, and comparison was made with the conventional diamond lattice approach. Statistical mechanical principles were used to relate the measured order parameters to the lipid bilayer structural parameters: the hydrocarbon thickness and the mean interfacial area per lipid. At fixed temperature, the area decreases with increasing acyl length, indicating increased van der Waals attraction for longer lipid chains. However, the main effect of increasing the acyl chain length is on the hydrocarbon thickness rather than on the area per lipid. Expansion coefficients of the structural parameters are reported and interpreted using an empirical free energy function that describes the force balance in fluid bilayers. At the same absolute temperature, the phosphatidylcholine (PC) series exhibits a universal chain packing profile that differs from that of phosphatidylethanolamines (PE). Hence, the lateral packing of phospholipids is more sensitive to the headgroup methylation than to the acyl chain length. A fit to the area per lipid for the PC series using the empirical free energy function shows that the PE area represents a limiting value for the packing of fluid acyl chains.     

Structure and interactions of fully hydrated dioleoylphosphatidylcholine bilayers.

This study focuses on dioleoylphosphatidylcholine (DOPC) bilayers near full hydration. Volumetric data and high-resolution synchrotron x-ray data are used in a method that compares DOPC with well determined gel phase dipalmitoylphosphatidylcholine (DPPC). The key structural quantity obtained is fully hydrated area/lipid A0 = 72.2 +/- 1.1 A2 at 30 degrees C, from which other quantities such as thickness of the bilayer are obtained. Data for samples over osmotic pressures from 0 to 56 atmospheres give an estimate for the area compressibility of KA = 188 dyn/cm. Obtaining the continuous scattering transform and electron density profiles requires correction for liquid crystal fluctuations. Quantitation of these fluctuations opens an experimental window on the fluctuation pressure, the primary repulsive interaction near full hydration. The fluctuation pressure decays exponentially with water spacing, in agreement with analytical results for soft confinement. However, the ratio of decay length lambda(fl) = 5.8 A to hydration pressure decay length lambda = 2.2 A is significantly larger than the value of 2 predicted by analytical theory and close to the ratio obtained in recent simulations. We also obtain the traditional osmotic pressure versus water spacing data. Our analysis of these data shows that estimates of the Hamaker parameter H and the bending modulus Kc are strongly coupled.     

Changes in phosphatidylcholine headgroup tilt and water order induced by monovalent salts: molecular dynamics simulations

The association between monovalent salts and neutral lipid bilayers is known to influence global bilayer structural properties such as headgroup conformational fluctuations and the dipole potential. The local influence of the ions, however, has been unknown due to limited structural resolution of experimental methods. Molecular dynamics simulations are used here to elucidate local structural rearrangements upon association of a series of monovalent Na(+) salts to a palmitoyl-oleoyl-phosphatidylcholine bilayer. We observe association of all ion types in the interfacial region. Larger anions, which are meant to rationalize data regarding a Hofmeister series of anions, bind more deeply within the bilayer than either Cl(-) or Na(+). Although the simulations are able to reproduce experimentally measured quantities, the analysis is focused on local properties currently invisible to experiments, which may be critical to biological systems. As such, for all ion types, including Cl(-), we show local ion-induced perturbations to headgroup tilt, the extent and direction of which is sensitive to ion charge and size. Additionally, we report salt-induced ordering of the water well beyond the interfacial region, which may be significant in terms of hydration repulsion between stacked bilayers.