立氏立克次体实时荧光定量聚合酶链反应检测方法的建立

目的采用TaqMan-MGB探针建立立氏立克次体实时荧光定量聚合酶链反应（PCR）检测方法.方法依据立氏立克次体外膜蛋白B基因序列设计引物和探针,以克隆的ompB基因片段为DNA模板,在ABI 7900型荧光定量PCR检测仪上建立实时荧光定量PCR检测方法.结果建立的定量标准曲线Ct值与模板拷贝数呈线性关系（R2=0.996）,最低检测浓度为5拷贝/μl;用荧光定量PCR方法检测其他相关立克次体和常见非立克次体病原菌,检出结果均为阴性.用该方法检测立氏立克次体感染的豚鼠血液标本、小鼠脾脏标本及细胞培养标本,检测的结果与立氏立克次体感染相关.结论研究建立的检测立氏立克次体实时荧光定量PCR方法具有高特异性和高敏感性,适用于快速检测各种样本中的立氏立克次体和立氏立克次体感染早期的实验室诊断.     

实时荧光定量PCR快速检测人鼻病毒

目的应用实时荧光定量PCR方法快速检测HRV,为HRV实验室早期诊断、HRV流行病监测和应急处理提供快速诊断手段。方法以HRV的5'UTR为检测靶基因,设计引物和探针,评价方法的特异性、灵敏性和重复性,并用该方法对384份临床呼吸道感染样本进行检测。结果以HRV、H1N1、Hco V、流感等病毒进行实时荧光定量PCR,只有HRV出现荧光信号,证明该法特异性强;以已知浓度HRV标准品稀释后进行实时荧光定量PCR,灵敏性达10 copies/μl,重复性试验Ct平均值变异系数0.58%~2.41%,384份临床样本中检测HRV阳性为21份,检测耗时比传统方法短。结论 HRV实时荧光定量PCR法有着快速、敏感、特异性强等优点,适用于HRV早期诊断、分子流行病学调查研究。     

重组酶介导扩增方法快速检测黄热病毒

目的本研究采用重组酶介导的等温核酸扩增方法(RAA),通过使用逆转录酶,建立黄热病毒的一步法等温核酸扩增(RT-RAA)方法。方法根据黄热病毒基因组保守序列设计引物和探针,建立并分析RT-RAA的重复性、特异性、灵敏度;以所建立方法对黄热病毒样本进行检测,同时以基因测序进行验证。结果黄热病毒RT-RAA扩增,体系中加入40 U的逆转录酶扩增效果最佳。该方法检测时间短(20 min),并且灵敏度高,检测下限可达100 copy,与登革病毒、西尼罗病毒、日本乙型脑炎病毒、基孔肯雅热病毒等蚊媒病毒无交叉反应,具有良好的特异性。结论构建的黄热病毒RT-RAA方法具有快速、特异以及灵敏的特点,适应于黄热病毒的口岸快速检测。     

多重实时荧光定量RT-PCR法快速检测3种蚊媒病毒

目的建立多重实时荧光定量RT-PCR快速检测方法,用于黄热病毒(YFV)、登革热病毒(DFV)、基孔肯雅热病毒(CHIKV)的同时检测和鉴别诊断。方法分别针对YFV 3'UTR基因、DFV 3'UTR C-M基因和CHIKV nsp2基因保守区设计特异性引物和Taq Man探针,建立并优化多重实时荧光定量RT-PCR反应体系,评价方法的特异性、敏感性和稳定性,并用临床标本进行验证。结果该方法可同时检测YFV、DFV和CHIKV,检测结果的特异性强,灵敏度达1×103copy/ml,结果的重复性很好,其变异系数分别为1.03%、0.90%和0.34%。临床确诊的11例DFV标本,5例CHIKV标本和10例黄热病疫苗标本核酸检测结果皆为阳性。结论研究建立的多重实时荧光定量RT-PCR方法可同时检测YFV、DFV和CHIKV,灵敏度高、特异性强、稳定性好,是一种可同时快速检测多种蚊媒病毒的新方法。     

普马拉病毒实时荧光定量RT-PCR方法的建立

目的建立一种适应国境口岸地区特定鼠种中普马拉病毒实时荧光定量RT-PCR检测方法。方法利用Beacon Designer7.0软件设计引物和探针,以人工合成普马拉病毒S基因的片段作为模板,进行实时荧光定量RT-PCR检测,并验证该方法的灵敏度及特异性。结果模板的Ct值与模板稀释浓度的对数存在良好的线性关系,标准曲线y=-3.122x＋38.605,R2=0.995,PCR扩增效率为109.1%,其最低检出限为31.6copies/μl。结论建立的实时荧光定量RT-PCR方法特异性好、灵敏度高,适合于普马拉病毒的快速检测。     

基孔肯雅病毒荧光定量PCR检测方法的建立

目的建立一种快速、敏感、特异的实时荧光定量PCR方法,检测基孔肯雅病毒。方法通过序列比对挑选出基孔肯雅病毒基因组中高度保守的序列,在此序列上设计引物及TaqMan探针,建立实时荧光定量PCR反应体系。结果经优化的荧光定量PCR方法有较好的灵敏度和特异性,对阳性对照质粒标准品的灵敏度可达21拷贝/μl,通过检测与传播媒介相似的流行性乙型脑炎病毒、黄热病毒、登革热病毒无交叉反应。结论该方法的建立在基孔肯雅热的疾病防控方面有较好的应用前景。     

实时荧光RT-PCR技术在黄热病快速检测中的应用

[目的]建立黄热病病毒的实时荧光RT-PCR检测方法并用于口岸黄热病的快速检测. [方法]体外转录黄热病病毒5-UTR的部分序列核酸作为阳性对照模板,设计实时荧光RT-PCR不同的反应程序和引物/探针浓度的组合,摸索最佳反应体系条件并用于黄热病疫苗和入境发热患者标本的检测.[结果]建立的黄热病病毒的实时荧光RT-PCR检测方法最佳反应体系为:2xRT-PCR缓冲液10 μl,正向引物5UTR-FP终浓度750 nM,反向引物5UTR-RP终浓度750 nM,探针5UTR-Probe终浓度500 nM,25xRT-PCR Enzyme Mix 0.8μl,RNA 5μl,用DEPC H2O补足到反应总体积20μl;Roche lightcycler仪器适用的扩增程序为:45℃10 min,95℃10 min;95℃5 s,56℃10 s(single),72℃15 s,45次循环.该方法最低检测限约为20copies病毒/反应,与登革病毒、西尼罗病毒、日本脑炎病毒、基孔肯雅病毒等蚊媒病毒无交叉反应.应用该方法对黄热病疫苗、媒介蚊虫和入境发热患者的血清标本进行检测,结果为黄热病疫苗核酸阳性,其他标本为阴性.[结论]该实时荧光RT-PER方法灵敏度高,特异性强,适用于黄热病病毒的快速检验,防止该病在国境口岸的传入.     

EV71TaqMan PCR检测方法的建立及初步应用

[目的]建立肠道病毒71型(enterovirus71)核酸的特异、快速、敏感的TaqMan探针PCR检测方法. [方法]根据GeneBank发表的EV71全基因组序列,在其VP1基因区段设计特异的引物与探针;以构建的重组质粒为模板,优化探针和引物浓度、最佳反应条件,建立最优化的实时定量PCR方法,评价其灵敏性、特异性和稳定性. [结果]引物和探针有良好的特异性,使用浓度为为0.8μM和0.4μM,可检测到100个RNA拷贝数,较传统的RT-PCR检测方法敏感性提高了100倍;同一样品Ct值重复检测3次,变异系数小于5%,表明该体系具有较好的稳定性. [结论]本研究建立的肠道病毒EV71-TaqMan荧光定量PCR方法,具有特异、灵敏、快速的特点,适用于由EV71感染引起的手足口病等的实验室早期诊断.     

实时荧光定量PCR检测普氏立克次体

目的 采用新型TaqMan—MGB探针建立检测普氏立克次体的实时荧光定量PCR方法。方法 根据普氏立克次体外膜蛋白B的基因（ompB）序列设计引物和探针，以克隆的ompB基因片段作DNA模板，在荧光定量PCR检测仪（ABI7900型）上建立实时荧光定量检测方法。结果 建立的定量标准曲线的循环阈值（Q）与模板拷贝数呈良好的线性关系（r=0．999）；与巢式PCR相比较，荧光定量PCR检测敏感性是其100倍。用荧光定量PCR检测莫氏立克次体及其他相关立克次体和细菌DNA，检出结果均为阴性。用荧光定量PCR检测普氏立克次体感染的豚鼠血标本，某些样本检测为阳性，而用巢式PCR检测的结果均为阴性。结论 研究中建立的检测普氏立克次体实时荧光定量PCR具有很高的特异性和敏感性，适合于快速检测样本中微量普氏立克次体DNA，可用作临床实验室快速确诊流行性斑疹伤寒。     

广州口岸入境外籍人员黄热病毒核酸及抗体水平分析

目的了解广州口岸入境外籍人员中黄热病毒核酸及该人群黄热病毒抗体水平,为研究黄热病输入我国的风险及口岸黄热病防控工作提供依据。方法对来自黄热病高风险区,离开高风险区6天,无症状且未能出示黄热病疫苗接种记录的外籍人,开展流行病学调查,进行黄热病毒核酸、抗体检测及数据分析。结果在2017-2019年间,结合流行病学调查随机抽选共839份目标人群的血清样本列入研究组,用荧光定量PCR法进行黄热病毒核酸检测,结果均为阴性;同时使用间接免疫荧光法(IFA)进行黄热病毒抗体IgG检测,结果显示阳性率为56.73%。其中年龄、人员类别和来源国是IgG抗体阳性率的影响因素。结论我国存在黄热病输入风险,对来自黄热病高风险区的人员开展黄热病毒核酸及抗体检测有助于口岸黄热病的防控工作;来自黄热病高风险区的外籍人员,黄热病毒抗体阳性率随年龄增加而升高。     

Calcium intake in the elderly: validation of a dietary questionnaire

Epidemiological studies of calcium and osteoporosis have been hampered by the lack of a suitable tool for assessing calcium intake. This report describes a new frequency and amount questionnaire for measuring present and past calcium intake in the elderly. The validity of the questionnaire was tested against two commonly used standards of dietary assessment, five-day duplicate diets and seven-day weighed dietary inventories. The resulting correlation coefficients were, respectively, r = 0.76 and r = 0.69, while that for repeatability was r = 0.84. Furthermore, the questionnaire categorized subjects into thirds of the distribution of intake with almost no gross misclassification. It is suggested that the present findings may be extended to the majority of normal, healthy elderly subjects, implying wide application for the questionnaire in the assessment of calcium intake in the elderly.     

The human drama of genetics: 'hard' and 'soft' media representations of inherited breast cancer

This paper examines media coverage of 'breast cancer genetics', and explores its implications for public understanding. We present a content analysis of coverage in British newspapers and look at a variety of popular forms, including women's magazines, television soap opera and radio drama. Genetic/inherited risk receives a great deal of coverage across a wide range of media formats and outlets. Much of this attention has focused on individuals from 'high risk families' and dilemmas around prophylactic mastectomies. Through examining media coverage, combined with interviews with media personnel and their sources, we show why this story proved so attractive to the media and highlight the different production values which influence coverage. Finally, we introduce preliminary findings from focus group discussions to demonstrate how such 'human interest' framing has engaged audience attention and influenced public understandings. The paper concludes by highlighting the implications for analysing, predicting, and engaging with, media representations of science.     

Worker education in the primary prevention of occupational dermatoses

This paper reports the evaluation of a skin care education programmeconducted on a fine chemicals manufacturing site where over1,000 employees are located. Approximately 60% are involvedin chemical manufacture. Over a 12 month period production staffreceived training in prevention of occupational dermatoses linkedto a site-wide poster initiative. The incidence of new casesof occupational dermatoses fell from 0.055 (70 cases in 1,277employees) to 0.021 (27 cases in 1,277 employees) before andafter the intervention respectively (p<0.0001). After otherfactors such as chemicals handled, observer bias and changesin reporting related to socioeconomic climate were taken intoaccount it is concluded that this study demonstrates the importanceof worker education as a tool for primary prevention of disease.Training materials such as video and poster presentations maybe effectively used in the chemical manufacturing industry asan adjunct to prevention and control of exposure to substanceshazardous to the skin. Such methods may also be used in otherindustries where there are significant risks of dermatoses.     

Regional Variation in Travel-related Illness acquired in Africa,March 1997–May 2011

To understand geographic variation in travel-related illness acquired in distinct African regions, we used the GeoSentinel Surveillance Network database to analyze records for 16,893 ill travelers returning from Africa over a 14-year period. Travelers to northern Africa most commonly reported gastrointestinal illnesses and dog bites. Febrile illnesses were more common in travelers returning from sub-Saharan countries. Eleven travelers died, 9 of malaria; these deaths occurred mainly among male business travelers to sub-Saharan Africa. The profile of illness varied substantially by region: malaria predominated in travelers returning from Central and Western Africa; schistosomiasis, strongyloidiasis, and dengue from Eastern and Western Africa; and loaisis from Central Africa. There were few reports of vaccine-preventable infections, HIV infection, and tuberculosis. Geographic profiling of illness acquired during travel to Africa guides targeted pretravel advice, expedites diagnosis in ill returning travelers, and may influence destination choices in tourism.     

Self-reported health problems among Swedish miners one year after unemployment

Unemployment is considered to be a public health concern sincedeterioration in the health of the unemployed is often anticipated.However, for some groups, such as miners, unemployment mightimprove health due to a cessation of potentially harmful occupationalexposures. This study evaluates the health of 79 miners in oneSwedish iron-ore mine, and 226 age-matched controls from thegeneral population, during one year after the closure of themine. The participants received a questionnaire regarding medicalhistory and subjective symptoms at the beginning of the studyperiod, and after one year. Statistically significant negativeeffects on self-reported health attributable to unemploymentwere not found, although neuropsychiatric symptoms were morecommon among the unemployed miners. The miners reported a statisticallysignificant improvement in grip force (p=0.031). They had asignificantly higher prevalence of symptoms associated withmining related exposures when compared with the population controls;pain in the upper extremities [relative risk (RR)=2.27, 95%confidence interval (Cl)=1.44–3.59), back pain (RR=1.84;Cl=1.237–2.75), vasospastic disease of the fingers (RR=2.05;Cl=1.18–3.57) and obstructive respiratory symptoms (attacksof dyspnea and wheezing: RR=3.67; Cl=1.167–11.6).     

Excès de cas humains et animaux de tularémie en France au cours de l’hiver 2007–2008 : émergence ou phénomène isolé ?

Context

Tularemia is a zoonosis affecting humans and hares in France. We describe the results of surveillance in both species, in 2007 and 2008.

Methods

Human tularemia cases are mandatorily notifiable in France since 2003. In hares, surveillance relies on volunteer hunter associations in all districts of the country. Data from mandatory reports and volunteer surveillance in 2007/2008 were analyzed and compared with previous results.

Results

In 2007/2008, 144 cases were reported in humans and 117 cases in hares. This was a 100% increase compared to previous years. Human cases differed from those of previous years only by the frequency of contact with breeding animals. Human cases without any documented risk exposure were also more frequent.

Conclusion

An increase of tularemia cases occurred in 2007/2008 in both species. Complementary studies are needed to identify the species reservoir in France to understand the causes of this peak of cases.     

深圳公立医院管理体制改革制度设计

深圳公立医院管理体制改革实行政事分开、管办分开,在理事会架构下按法定机构模式组建市医管中心,落实公立医院运营管理自主权.作者从当前公立医院管理体制的弊端入手,介绍了深圳市进行公立医院管理体制改革的基本思路及改革方案设计的主要举措,深入剖析了的改革方案的特点,并对改革效果进行了预测.     

Effects of nitrogen dioxide on red blood cells of rats: Changes in components of red cell membranes during in vivo exposure to NO2

Red cell membranes, prepared from red blood cells of rats exposed to 4, 10, or 20 ppm nitrogen dioxide (NO₂) for 1 to 10 days, were examined for evidence of changes in membrane components. Appreciable changes were not found in contents of phospholipid and cholesterol during exposure to 10 ppm NO₂. By contrast, protein content altered with the time of exposure. Moreover, changes in protein composition were observed by employing sodium dodecyl sulfate — polyacrylamide gel electrophoresis. Twenty-four-hour exposure to NO₂ at the concentration above 10 ppm resulted in a marked increase in the percentage of lysophosphatidylethanolamine (LysoPE) to the total phospholipids. The prolonged exposure to 10 ppm NO₂ gave rise to a further increase in LysoPE, whereas the percentage of phosphatidylethanolamine (PE) showed a gradual decrease. A 1-day exposure to 4.0 ppm NO₂ also caused an increase in sialic acid content and decreases in those of PE and hexose. In addition to contents of these components the percentage of LysoPE increased 5 days after exposure and the elevated values were maintained up to the end of exposure period. These results demonstrate that red blood cells in circulation exhibit different membrane properties in terms of lipid and carbohydrate composition during 10 days of exposure to 4.0 ppm NO₂.     

The design of hazard risk assessment matrices for ranking occupational health risks and their application in mining and minerals processing

Two hazard risk assessment matrices for the ranking of occupational health risks are described. The qualitative matrix uses qualitative measures of probability and consequence to determine risk assessment codes for hazard-disease combinations. A walk-through survey of an underground metalliferous mine and concentrator is used to demonstrate how the qualitative matrix can be applied to determine priorities for the control of occupational health hazards. The semi-quantitative matrix uses attributable risk as a quantitative measure of probability and uses qualitative measures of consequence. A practical application of this matrix is the determination of occupational health priorities using existing epidemiological studies. Calculated attributable risks from epidemiological studies of hazard-disease combinations in mining and minerals processing are used as examples. These historic response data do not reflect the risks associated with current exposures. A method using current exposure data, known exposure-response relationships and the semi-quantitative matrix is proposed for more accurate and current risk rankings.