大鼠初级感觉神经元P2X3受体的表达及其与SP的关系

目的研究在大鼠初级感觉神经元细胞上P2X3受体的表达情况及其与P物质的关系。方法取SD大鼠背根神经节(DRG)和三叉神经节(TG)固定后切片;用抗P2X3受体抗体和抗SP抗体进行免疫组织化学反应,并通过两种不同的显色方法同时进行P2X3受体和SP的双标。结果P2X3免疫反应阳性细胞主要集中在小细胞和中等细胞(其中在TG,P2X3-ir阳性神经元约占整个细胞的24.8%;在DRG约31.7%的神经元是P2X3-ir阳性),并且在DRG和TG细胞上均存在有P2X3受体和SP共存(TG上的双标细胞占P2X3-ir阳性细胞总数的36.26%,DRG上占46.81%)。结论由于ATP门控阳离子通道受体P2X3本身就与伤害性感受的初级传入有关,而它与SP的共存可提示当组织中的ATP释放时可以通过P2X3受体作用于含SP的伤害性感觉神经末梢上,促使SP释放引起痛觉过敏。     

The use of intensity‐based Doppler variance method for single vessel response to functional neurovascular activation

This study presents 1 use of optical coherence tomography (OCT) angiography technique to examine neurovascular coupling effect. Repeated B‐scans OCT recording is performed on the rat somatosensory cortex with cranial window preparation while its contralateral forepaw is electrically stimulated to activate the neurons in rest. We use an intensity‐based Doppler variance (IBDV) algorithm mapped cerebral blood vessels in the cortex, and the temporal alteration in blood perfusion during neurovascular activation is analyzed using the proposed IBDV quantitative parameters. By using principal component analysis‐based Fuzzy C Means clustering method, the stimulus‐evoked vasomotion patterns were classified into 3 categories. We found that the response time of small vessels (resting diameter 14.9 ±6.6 μm), middle vessels (resting diameter 21.1 ±7.9 μm) and large vessels (resting diameter 50.7 ±6.5 μm) to achieve 5% change of vascular dilation after stimulation was 1.5, 2 and 5.5 seconds, respectively. Approximately 5% peak change of relative blood flow (RBF) in both small and middle vessels was observed. The large vessels react slowly and their responses nearly 4 seconds delayed, but no significant change in RBF of the large vessels was seen.      

A novel carbonyl reductase from Pichia stipitis for the production of ethyl (S)-4-chloro-3-hydroxybutanoate

An NADPH-dependent carbonyl reductase (PsCR) gene from Pichia stipitis was cloned. It contains an open reading frame of 849 bp encoding 283 amino acids whose sequence had less than 60% identity to known reductases that produce ethyl (S)-4-chloro-3-hydroxybutanoates (S-CHBE). When expressed in Escherichia coli, the recombinant PsCR exhibited an activity of 27 U/mg using ethyl 4-chloro-3-oxobutanoate (COBE) as a substrate. Reduction of COBE to (S)-CHBE by transformants in an aqueous mono-phase system for 18 h, gave a molar yield of 94% and an optical purity of the (S)-isomer of more than 99% enantiomeric excess.     

Intact Flagellar Motor of Borrelia burgdorferi Revealed by Cryo-Electron Tomography: Evidence for Stator Ring Curvature and Rotor/C-Ring Assembly Flexion

The bacterial flagellar motor is a remarkable nanomachine that provides motility through flagellar rotation. Prior structural studies have revealed the stunning complexity of the purified rotor and C-ring assemblies from flagellar motors. In this study, we used high-throughput cryo-electron tomography and image analysis of intact Borrelia burgdorferi to produce a three-dimensional (3-D) model of the in situ flagellar motor without imposing rotational symmetry. Structural details of B. burgdorferi, including a layer of outer surface proteins, were clearly visible in the resulting 3-D reconstructions. By averaging the 3-D images of ∼1,280 flagellar motors, a ∼3.5-nm-resolution model of the stator and rotor structures was obtained. flgI transposon mutants lacked a torus-shaped structure attached to the flagellar rod, establishing the structural location of the spirochetal P ring. Treatment of intact organisms with the nonionic detergent NP-40 resulted in dissolution of the outermost portion of the motor structure and the C ring, providing insight into the in situ arrangement of the stator and rotor structures. Structural elements associated with the stator followed the curvature of the cytoplasmic membrane. The rotor and the C ring also exhibited angular flexion, resulting in a slight narrowing of both structures in the direction perpendicular to the cell axis. These results indicate an inherent flexibility in the rotor-stator interaction. The FliG switching and energizing component likely provides much of the flexibility needed to maintain the interaction between the curved stator and the relatively symmetrical rotor/C-ring assembly during flagellar rotation.Flagellum-based motility plays a critical role in the biology and pathogenesis of many bacteria (3, 6, 17, 31). The well-conserved flagellum is commonly divided into three physical parts: the flagellar motor, the helically shaped flagellar filament, and the hook which provides a universal joint between the motor and the filament. In most bacteria, counterclockwise rotation of the flagella results in bundling of the helical flagella and propulsion of the cell through liquid or viscous environments. Clockwise rotation of the flagellar motor results in random turning of the cell with little translational motion (“tumbling”). Bacterial motility is thus a zigzag pattern of runs and tumbles, in which chemotactic signals favor running toward attractants and away from repellents (3).Borrelia burgdorferi and other closely related spirochetes are the causative agents of Lyme disease, which is transmitted to humans via infected Ixodes ticks (40). Spirochetes have a distinctive morphology in that the flagella are enclosed within the outer membrane sheath and are thus called periplasmic flagella (6). The flagellar motors are located at both ends of the cell and are coordinated to rotate in opposite directions during translational motion and in the same direction (i.e., both clockwise or both counterclockwise) during the spirochete equivalent of tumbling, called “flexing” (6, 15). Spirochetes are also capable of reversing translational motion by coordinated reversal of the direction of motor rotation at both ends of the cell. Rotation of the flagella causes a serpentine movement of the entire cell body, allowing B. burgdorferi to efficiently bore its way through tissue and disseminate throughout the mammalian host, resulting in manifestations in the joints, nervous system, and heart (40).The flagellar motor is an extraordinary nanomachine powered by the electrochemical potential of specific ions across the cytoplasmic membrane (3). Current knowledge of the flagellar motor structure and rotational mechanisms is based primarily on studies of Escherichia coli and Salmonella enterica and is summarized in several recent comprehensive reviews (3, 22, 31, 39, 42). The flagellar motor is constructed from at least 20 different kinds of proteins. The approximate location of these flagellar proteins has been determined by a variety of approaches and appears to be relatively consistent in a wide variety of bacteria. It can be divided into several morphological domains: the MS ring (FliF, the base for the flagellar motor); the C ring (FliG, FliM, and FliN, the switch complex regulating motor rotation); the export apparatus (multiple-protein complex located at the cytoplasmic side of the MS ring); the rod (connecting the MS ring and the hook); the L and P rings on the rod (thought to serve as bushings at the outer membrane and at the peptidoglycan layer, respectively); and the stator, which is the motor force generator embedded in the cytoplasmic membrane. Electron microscopy studies of the purified flagellar motor have provided a detailed view of the rotor/C-ring assembly (11, 44). However, there is no structural information on the stator and the export apparatus in these reconstructions, because these membrane-associated structures are not retained following detergent extraction during the extensive basal body purification process. The stator and the export apparatus were visualized by using freeze fracture preparations of cytoplasmic membranes. It appears that 10 to 16 stator units form circular arrays in the membrane (9, 20). Part of the export apparatus is located in the central space of the C ring (18). Recently a 7-nm-resolution structure of the intact flagellar motor in situ was revealed by averaging 20 structures obtained using cryo-electron tomography (cryo-ET) of Treponema primitia cells (32). Further analysis of the intact flagellar motor structure would lead to a better understanding of the motor protein distribution, the rotor-stator interaction, and the mechanism of bacterial motility.Cryo-ET has emerged as a three-dimensional (3-D) imaging technique to bridge the information gap between X-ray crystallographic and optical microscopic methods (24, 30). This process involves rapidly freezing viable cells, collecting a series of electron micrographs at different angles, and computationally combining the resulting images into a 3-D density map. Cryo-ET allows investigation of the structure-function relationship of molecular complexes and supramolecular assemblies in their cellular environments without fixation, dehydration, embedding, or sectioning artifacts. Spirochetes are well suited for cryo-ET analysis because of their narrow cell diameter (typically 0.2 to 0.3 μm). Recently the cellular architecture of Treponema primitia, Treponema denticola, and B. burgdorferi, as well as the configuration of the B. burgdorferi periplasmic flagella, were revealed by cryo-ET (7, 16, 26, 33). In combination with advanced computational methods, cryo-ET is currently the most promising approach for determining the cellular architecture in situ at molecular resolution (30). We have developed novel strategies for capturing and averaging thousands of 3-D images of large macromolecular assemblies to obtain ∼2.0-nm-resolution structures (28, 29).In this study, we present the molecular structures of infectious wild-type (WT) and mutant B. burgdorferi organisms and their flagellar motors in situ using high-throughput cryo-ET and 3-D image analysis. By averaging subvolumes of 1,280 flagellar motors from 322 cells, we obtained a ∼3.5-nm-resolution model of the intact flagellar motor, providing a detailed view of rotor-stator interactions. In addition, detergent treatment of intact cells provided a preliminary identification of the rotor and stator structures. Through the comparison of WT and mutant cells, we have also determined the location of the flgI gene product in the B. burgdorferi flagellar motor.     

几种杀菌剂对黄瓜疫霉菌的毒力测定

采用菌丝生长速率法检测60%嘧菌酯悬浮剂、80%代森锰锌可湿性粉剂、75%百菌清可湿性粉剂、60%唑醚·代森联水分散粒剂、2%蛇床子素微乳剂、64%杀毒矾可湿性粉剂等6种杀菌剂对黄瓜疫霉菌的室内毒力。结果表明,唑醚·代森联和蛇床子素对黄瓜疫霉菌的毒力最高,EC50值分别为3.4143 mg·L-1和10.4641 mg·L-1。将两种原药混配后,表现出增效作用,在两者混配比为1∶2时增效最明显,EC50值为5.8178 mg·L-1。     

未来气候变化对河南省冬小麦需水量和缺水量的影响预估

采用美国农业部土壤保持局推荐的方法计算有效降水量,应用Penman-Monteith模型和作物系数法计算需水量,在对河南省1981—2010年冬小麦生育期内有效降水量、需水量和缺水量分析的基础上,结合《排放情景特别报告》的两种排放情景A2(强调经济发展)和B2(强调可持续发展)预估的未来气候情景,探讨了未来气候情景下河南省冬小麦的有效降水量、需水量和缺水量的时空演变规律及其主要气候影响因素.结果表明: 从整体上看,相对于基准时段(1981—2010年),A2和B2情景下,不同时段冬小麦全生育期的有效降水量、需水量和缺水量均表现出增加趋势,有效降水量均以2030s时段增加最多,分别增加33.5%和39.2%;需水量均以2010s时段增加最多,分别增加22.5%和17.5%,年代间呈现明显递减趋势;缺水量在A2情景下以2010s时段增加(23.6%)最多,B2情景下以2020s时段增加(13.0%)最多.偏相关分析表明,A2和B2情景下,太阳总辐射是影响河南省冬小麦需水量和缺水量变化的主要气候因素.由于地理环境和气候条件的差异,不同时段河南省冬小麦全生育期有效降水量、需水量和缺水量的距平百分率在空间分布上具有差异.未来河南省水资源可能更趋于短缺.     

污染预测的非线性参数模型

本文探索了Cr~（3 ）,Cr~（6 ）溶液浓度与绿豆幼苗根过氧化物酶活性之间的相关关系,建立了线性与非线性的数学模型,经相关指数,剩余标准差检验,找出了拟合度比较高的非线性参数模型．本文的结果为利用过氧化物酶活性增长率作为水环境受重金属污染的生物监测指标提供了理论依据．     

海南省香蕉枯萎病病原菌的分离鉴定及1号、4号小种的生物学特性

香蕉枯萎病是目前国内和世界很多地区香蕉种植区的一种毁灭性病害,目前尚未找到有效的防治方法.本文采集了海南省不同地区香蕉枯萎病的病原菌,通过经典病理学与分子生物学的方法进行了鉴定,此外还优化了快速检测的方法.结果表明,通过接种巴西香蕉和粉蕉进行致病性检测以及分离菌rDNA-ITS测序结果,明确鉴定出供试的香蕉分离菌株为4号生理小种,粉蕉分离菌株为1号生理小种.为了对1号、4号生理小种有更多的了解,本文还对1号、4号小种菌株生物学特性进行了测定,确定了影响菌丝生长的最佳条件.结果显示,pH 5时最适合1号、4号生理小种的菌丝生长,适合菌丝生长的温度为19～28℃之间,温度达到37℃时菌丝不能生长,碳源对菌丝的生长影响不大.这些方法和数据可以有利于田间香蕉枯萎病病害的检测和最终确定,生物学特性的测定可以为耕作技术的改进和农药的研发提供参考依据.     

群体遗传结构中的基因流

群体遗传结构上的差异是遗传多样性的一种重要体现,对群体遗传结构的研究已有较久的历史,而其中的基因流研究近些年来越来越受到重视。它对群体遗传学、进化生物学、保护生物学、生态学有着极其重要的作用。虽然传统的群体遗传学能估测基因流大小,但它的精确性还有很大局限性。随着生物技术的进步,对基因流的研究逐渐向分子水平过渡,应用蛋白质电泳技术、分子标记技术(RAPD、RFLP、VNTR、ISSR、DNA测序等)方法对群体间基因流的流动水平进行了深入细致的研究。通过综述群体遗传结构的几种模式:陆岛模式、海岛模式、阶石模式、距离隔离模式、层次模式,以及在群体遗传结构的几种模式基础上的基因流的研究方法、作用、地位和近些年来研究者的研究成果,并指出了这些方法的局限性。