多重聚合酶链反应快速鉴别葡萄球菌及mecA基因检测

目的：建立快速鉴定葡萄球菌及检测mecA基因的多重聚合酶链反应（PCR）诊断方法。方法：以葡萄球菌16SrRNA、金黄色葡萄球菌的nuc基因以及耐甲氧西林葡萄球菌（MRS）的mecA基因合成3对引物，采用多重PCR扩增法，快速鉴定葡萄球菌，同时判断MRS菌株。结果：107株葡萄球菌的16SrRNA基因扩增片段全部为阳性．与常规鉴定结果符合率为100％；nuc基因阳性48株。阴性59株，与金黄色葡萄球菌核酸酶试验一致；以PBP2a乳胶凝集试验结果为金标准，苯唑西林（含2％NaClMHA）及头孢西丁纸片扩散法的敏感性为98．7％和100％，特异性分别为75．0％和82．1％：PCR扩增法的敏感性和特异性均为100％。结论：多重PCR法能快速准确地从临床分离菌株中鉴别葡萄球菌，并可同步对mecA基因进行准确检测。     

16S rRNA及18S rRNA同时检测脑脊液中的病原菌

目的 提高脑脊液中病原菌的快速、准确鉴定率。方法 以细菌的保守序列16SrRNA和真菌的18SrRNA为通用引物，用多重PCR技术对标准菌株及46例中枢神经系统感染患者的脑脊液进行扩增，同时与脑脊液的培养结果进行比较。结果 标准菌株金黄色葡萄球菌、大肠埃希菌、脑膜炎奈瑟菌、结核分枝杆菌、化脓性链球菌及白色念球菌均出现特异条带，人白细胞DNA为阴性。46份脑脊液标本培养阳性39例，另有2例PCR阳性而培养阴性，后经诱导传代培养证实为L型金黄色葡萄球菌。PCR阳性41例，与培养分离鉴定结果符合率为100％。结论 多重PCR可快速鉴定脑脊液中病原菌，为临床及时治疗提供必要的诊断信息。     

宏基因组二代测序在重症风湿病合并感染诊断中的应用

目的 探讨宏基因组二代测序(mNGS)在重症风湿病合并感染诊断中的应用价值。方法 重症风湿病合并感染患者28例行mNGS检测和传统实验室病原学检测，分析两者检测结果及mNGS检测结果对治疗策略的影响。结果 mNGS共检测出33种细菌属、4种真菌属和17种病毒。重症风湿病合并感染患者mNGS检测阳性率高于传统实验室病原学检测阳性率(67.86%vs.21.43%)(P<0.05)。根据mNGS检测结果，18例临床治疗策略做了改变。25例好转出院，3例死亡。结论 mNGS可有效辅助重症风湿病合并感染患者的诊断。     

尿细菌培养与尿沉渣细菌定量计数对于泌尿系统感染的诊断比较

目的探讨尿沉渣细菌定量计数在泌尿系感染的诊断价值。方法以我院门诊拟诊尿路感染的120例患者为研究对象,收集所选患者的中断尿,分别采用尿沉渣定量检测与尿细菌培养,比较两组检测方法的结果,以尿细菌培养为金标准进行分析。结果所选120例标本中,尿细菌培养检测阳性77例(64.2%),尿沉渣定量计数测定阳性73例(60.8%),两种检测方法的阳性率比较,差异不显著无统计学意义(P>0.05)。尿沉渣细菌定量计数与尿细菌培养的阳性符合率为90.4%,阴性符合率为83.7%。结论与尿细菌培养检测结果比较,尿沉渣定量分析与其阳性率相一致,准确性较高,有一定的临床价值,能作为泌尿系感染快速筛查的检测方法,值得临床推广应用。     

小儿急性肠套叠细菌移位及IL-11变化的临床意义

目的探讨小儿急性肠套叠细菌移位、白细胞介素11（IL-11）的变化及临床意义。方法应用聚合酶链反应（PCR）定性检测肠套叠组及对照组全血细菌DNA;应用酶联免疫吸附试验（ELISA）检测肠套叠组术前、术后及对照组血清中IL-11浓度。结果正常对照组全血16SrRNA、大肠杆菌BG未检出,空气灌肠复位组16SrRNA阳性率30%,BG阳性率20%;手法复位组16SrRNA阳性率50%,BG阳性率60%;肠坏死肠切除组16SrRNA阳性率60%,BG阳性率70%;肠套叠患儿血清IL-11含量3组中升高有统计学意义,组间存在显著性差异。结论小儿急性肠套叠应用PCR技术可早期检测细菌移位,而IL-11的检测对于急性肠套叠细菌移位的辅助诊断及判断愈后有着重要的意义。     

鲍曼不动杆菌老年患者分离株中发现16S rRNA甲基化酶基因新亚型

目的了解鲍曼不动杆菌老年患者分离株16SrRNA甲基化酶和氨基糖苷类修饰酶基因型。方法采用PCR检测20株鲍曼不动杆菌老年患者分离株12种16SrRNA甲基化酶和氨基糖苷类修饰酶基因。结果7株检出16SrRNA甲基化酶基因（armA），19株检出氨基糖苷类修饰酶基因[其中aac（3）-I阳性率为95％，ant（3″）-I为95％，aac（3）-II为40％，aac（δ′）-Ib为15％）]。6号株armA基因测得序列翻译成氨基酸序列与美国核酸库（GenBank）已登录的armA氨基酸不同，为新亚型。结论本组鲍曼不动杆菌老年患者分离株95％携带16SrRNA甲基化酶和氨基糖苷类修饰酶基因。     

重症手足口病合并细菌感染的细菌培养分析

目的提示重症手足口病易发生呼吸道合并感染。方法分析我院重症手足口病患儿细菌培养送检结果。结果脑脊液培养无阳性,血培养阳性率为8.9%,痰培养阳性率高达50%,主要是条件致病菌感染。结论重症手足口患儿细胞免疫和体液免疫功能下降,呼吸系统黏膜感染防御能力降低,极易并发呼吸道细菌感染。     

荧光定量聚合酶链反应在小儿巨细胞病毒感染检测中的应用

目的　探讨荧光定量聚合酶链反应 (FQ- PCR)法检测小儿巨细胞病毒感染效果及临床意义。方法　应用 FQ- PCR法检测患儿血标本 HCMV- DNA。结果　 77例患儿血标本 FQ - PCR检测 HCMV - DNA阳性 19例 ,阳性率 2 4 .7% ,阳性标本的平均拷贝数为 1.16× 10 4 。结论　 FQ- PCR法对于 HCMV感染的早期诊断和疗效判断 ,具有重要意义。     

培养法检测肺炎支原体的临床应用及结果分析

目的探讨培养法检测肺炎支原体（MP）的临床应用价值。了解该地区儿童MP的感染情况。方法取患者咽拭子样本接种于MP选择鉴别培养基进行培养，并将检测结果与酶免法、PCR法检测结果进行比较分析。结果培养法检测945份样本，阳性率为37．99％（359／945）；ELISA法检测270例，MP-IgM阳性率为13．3％（36／270）。荧光定量PCR法与培养法阳性符合率为100％。结论培养法检测MP阳性明显高于ELISA法，可作为早期诊断的金标准，且方法简便快速，易于推广。     

应用PCR方法直接检测痰中耐甲氧西林葡萄球菌

目的建立直接用痰标本通过PCR方法快速检测耐甲氧西林葡萄球菌的方法。方法利用聚合酶链反应技术（PCR）快速检出耐甲氧西林葡萄球菌，建立一种从痰中快速提取DNA方法，以粗提DNA作为PCR模板，检测编码耐甲氧西林葡萄球菌青霉素结合蛋白2a（PBP2a）的mecA基因和细菌中均有的16SrRNA基因。结果364份临床痰标本经PCR方法检出mecA基因的38份，药敏法检出耐甲氧西林的葡萄球菌33份；38份mecA基因阳性的痰标本中有31份药敏法检测为耐甲氧西林的葡萄球菌，药敏法检出耐甲氧西林的葡萄球菌33份痰标本中有2份mecA基因阴性。16SrRNA基因片段在PCR中作为内部对照避免了假阴性结果的出现。结论用PCR直接检测痰中的mecA基因，是判断痰中是否含有耐甲氧西林的葡萄球菌的一种快速有效的方法。     

Efficacy of gut lavage,hemodialysis, and hemoperfusion in the therapy of paraquat or diquat intoxication

Clinical and in vitro investigations were carried out to test the efficacy of gut lavage, hemodialysis, and hemoperfusion in the treatment of poisoning with paraquat or diquat. In a patient suffering from diquat intoxication 130 times more diquat was removed by gut lavage 30 h after ingestion than was removed by complete aspiration of the gastric contents.Determination of in vitro clearances for paraquat and diquat by hemodialysis showed that, at serum concentrations of 1–2 ppm, such as are frequently encountered in poisoning in man, toxicologically relevant quantities of herbicide cannot be removed from the body. At a concentration of 20 ppm, on the other hand, hemodialysis proved to be effective, the clearance being 70 ml/min at a blood flow rate of 100 ml/min. The efficacy of hemoperfusion with coated activated charcoal was on the whole better. Especially at concentrations around 1–2 ppm, the clearance values for hemoperfusion were some 5–7 times higher than those for hemodialysis.In a patient suffering from paraquat poisoning, both hemodialysis as well as hemoperfusion were carried out. The in vitro results could be confirmed: At serum concentrations of paraquat less than 1 ppm no clearance could be obtained by hemodialysis while by hemoperfusion with activated charcoal quite high clearance values were measured and the serum level dropped down to zero.

---

Zusammenfassung Klinische Untersuchungen und Laboratoriumsversuche wurden durchgeführt, um die Wirksamkeit von Darmspülung, Hämodialyse und Hämoperfusion bei Paraquat- und Deiquat-Vergiftungen zu prüfen.Bei einem Patienten wurde 30 Std nach Deiquat-Aufnahme durch Darmspülung 130mal mehr Deiquat entfernt als durch vollständige Aspiration des Mageninhaltes. In vitro-Versuche ergaben, daß bei Blutserumkonzentrationen von 1–2 ppm, die bei Vergiftungen oft gemessen werden, durch Hämodialyse keine toxikologisch relevanten Paraquat- oder Deiquat-Mengen entfernt werden können. Dagegen erwies sich die Hämodialyse bei 20 ppm und einer Blutumlaufgeschwindigkeit von 100 ml/min mit einer Clearance von 70 ml/min als wirksam. Die Hämoperfusion mit beschicheter Aktivkohle war in diesen Versuchen aber eindeutig überlegen, denn insbesondere bei Konzentrationen um 1–2 ppm waren die Clearance-Werte 5–7mal höher als bei der Hämodialyse.Die in vitro-Ergebnisse wurden bei einem Patienten mit einer Paraquat-Vergiftung bestätigt: Bei Konzentrationen unter 1 ppm war die Hämodialyse wirkungslos, während durch Hämoperfusion relativ hohe Clearance-Werte erreicht wurden, so daß der Serumspiegel rasch unter die Nachweisgrenze abfiel.

     

In vitro studies on sterically stabilized liposomes (SL) as enzyme carriers in organophosphorus (OP) antagonism

This study describes a new approach for organophosphorous (OP) antidotal treatment by encapsulating an OP hydrolyzing enzyme, OPA anhydrolase (OPAA), within sterically stabilized liposomes. The recombinant OPAA enzyme was derived from Alteromonas strain JD6. It has broad substrate specificity to a wide range of OP compounds: DFP and the nerve agents, soman and sarin. Liposomes encapsulating OPAA (SL)* were made by mechanical dispersion method. Hydrolysis of DFP by (SL)* was measured by following an increase of fluoride ion concentration using a fluoride ion selective electrode. OPAA entrapped in the carrier liposomes rapidly hydrolyze DFP, with the rate of DFP hydrolysis directly proportional to the amount of (SL)* added to the solution. Liposomal carriers containing no enzyme did not hydrolyze DFP. The reaction was linear and the rate of hydrolysis was first order in the substrate. This enzyme carrier system serves as a biodegradable protective environment for the recombinant OP-metabolizing enzyme, OPAA, resulting in prolongation of enzymatic concentration in the body. These studies suggest that the protection of OP intoxication can be strikingly enhanced by adding OPAA encapsulated within (SL)* to pralidoxime and atropine.     

Aquatic Insects and Trace Metals: Bioavailability,Bioaccumulation, and Toxicity

Abstract

The uptake of metals from food and water sources by insects is thought to be additive. For a given metal, the proportions taken up from water and food will depend both on the bioavailable concentration of the metal associated with each source and the mechanism and rate by which the metal enters the insect. Attempts to correlate insect trace metal concentrations with the trophic level of insects should be made with a knowledge of the feeding relationships of the individual taxa concerned. Pathways for the uptake of essential metals, such as copper and zinc, exist at the cellular level, and other nonessential metals, such as cadmium, also appear to enter via these routes. Within cells, trace metals can be bound to proteins or stored in granules. The internal distribution of metals among body tissues is very heterogeneous, and distribution patterns tend to be both metal and taxon specific. Trace metals associated with insects can be both bound on the surface of their chitinous exoskeleton and incorporated into body tissues. The quantities of trace meals accumulated by an individual reflect the net balance between the rate of metal influx from both dissolved and particulate sources and the rate of metal efflux from the organism. The toxicity of metals has been demonstrated at all levels of biological organization: cell, tissue, individual, population, and community. Much of the literature pertaining to the toxic effects of metals on aquatic insects is based on laboratory observations and, as such, it is difficult to extrapolate the data to insects in nature. The few experimental studies in nature suggest that trace metal contaminants can affect both the distribution and the abundance of aquatic insects. Insects have a largely unexploited potential as biomonitors of metal contamination in nature. A better understanding of the physico-chemical and biological mechanisms mediating trace metal bioavailability and exchange will facilitate the development of general predictive models relating trace metal concentrations in insects to those in their environment. Such models will facilitate the use of insects as contaminant biomonitors.     

Alzheimer’s disease – current trends in basic and clinical research. Highlights from the 16th ECNP

Advances in the molecular biological knowledge of neuronal nicotinic acetylcholine receptors (nAChRs) have led to a growing interest by the pharmaceutical industry in the development of novel compounds that selectively modulate nAChR function. The ability of (-)-nicotine, an activator of nAChRs, to enhance attentional aspects of cognition in animals and humans, to exert neuroprotective and anxiolytic-like effects, and presumably to mediate the negative correlation between smoking and Alzheimer's (and Parkinson's) Disease, has focused interest on the potential therapeutic utility of modulators of nAChR function for treatment of some of the deficits associated with these progressive, neurodegenerative conditions. Numerous compounds are known which activate nAChRs and which might serve as lead compounds toward the development of such agents. The pharmacologic diversity of neuronal nAChR subtypes suggests the possibility of developing selective compounds which would have more favourable side-effect profiles than existing agents. This broader class of agents, collectively called cholinergic channel modulators (ChCMs), is anticipated to encompass compounds which would have more favourable side-effect profiles than existing agents, which generally exhibit low selectivity. This selectivity may be achieved by preferentially activating some subtypes of nAChRs (i.e., Cholinergic Channel Activators, ChCAs) or inhibiting the function of other subtypes (Cholinergic Channel Inhibitors, ChCIs). An overview of the biology of nAChRs and the rationale for the use of ChCMs for the treatment of dementia related to neurodegenerative diseases are presented, followed by a discussion of lead compounds and compounds under consideration for clinical evaluation.     

Steroidal sapogenin contents in some domestic plants

In order to find out the values of the steroid resources for the future use. the compositions and contents of steroidal sapogenins from 13 domestic plants have been investigated. As a result,Dioscorea nipponica, D. quinqueloba andSmilax china were found to have large amount of diosgenin. And pennogenin inTrillium kamtschaticum andParis verticillata, yuccagenin inAllium fistulosum, hecogenin inAgave americana and neochlorogenin inSolanum nigum were appeared to be major steroidal sapogenins.