手足口病病原体实时荧光RT-PCR反应体系的建立与临床应用

目的 建立一种快速、准确、特异件高的方法 检测手足口病病原体.方法 根据引起手足口病常见致病原肠道病毒71(EV71)、柯萨奇病毒A16(CA16)及肠道病毒(EV),设计相应的引物、探针对35例临床诊断的手足口病患儿及20例正常健康婴儿的粪便进行EV、EV71、CA16三种病原体实时荧光RT-PCR检测,同时对55分标本进行EV71病毒培养分离.结果 35例临床诊断手足口病患儿粪便中EV全部阳性,EV71阳性25例,CA16阳性8例,其中3例为EV71、CA16同时阳性,与临床诊断符合率为85.71%,20例健康体检婴儿粪便中EV病毒5例阳性,EV71、CA16均阴性.结论 荧光RT-PCR对手足口病病原体检测准确性高、特异性强,具有快速、廉价等特点,适合于手足口病的早期诊断.     

实时RT-PCR检测159例手足口病患儿样本中肠道病毒的结果分析

目的 了解2009年4-8月首都儿科研究所附属儿童医院手足口病患儿肠道病毒的感染状况,为临床诊治提供参考.方法 采集首诊手足口病159例患儿的咽拭子和疱疹液标本,以肠道病毒(EV)通用型、柯萨奇病毒A16(CA16)型、肠道病毒71(EV71)型核酸检测试剂盒,应用实时RT-PCR法检测标本中的肠道病毒.选取阳性标本扩增VP1区,产物进行序列测定和分析.结果 (1)EV、CA16、EV71的阳性病例数分别为152、102、43;阳性率为95.6%、64.2%、27.0%.(2)CA16占EV阳性的67.3%,EV71占EV阳性的28.3%,非CA16和EV71的EV病例7例,占EV阳性的4.6%.CA16:EV71为2.37:1.(3)部分阳性标本经测序验证与此法结果一致.结论 2009年我院手足口病患儿以EV71和CA16感染为主,EV71感染的手足口病比例较2007年出现明显上升.     

四重荧光RT-PCR检测手足口肠道病毒方法的研究

本文采用TaqMan探针技术,建立了含有非竞争性内标检测肠道病毒通用型并能对EV71型和CA16型进行分型的多重荧光RT-PCR检测方法,内标同步参与样品核酸的提取,不仅能够有效地监控样本中的抑制物,还能避免操作误差所造成的假阴性.     

2009年夏季174例手足口病患儿病原分析

目的分析本院2009年夏季手足口病患儿病原及其血清型特征,为临床早期诊断和疾病防控提供实验依据。方法2009年4—9月,首都儿科研究所附属儿童医院门诊就诊的174例手足口病患儿咽拭子和疱疹液,实时荧光RT—PCR方法检测肠道病毒通用型（EV）、柯萨奇A16型（CA16）、肠道病毒71型（EV71）。131例患儿双份血清,同时检测CA16、EV71IgM抗体。CA16和EV71阳性标本扩增VP1区基因片段后测序,进行同源性和系统进化分析。结果（1）EV、CA16、EV71阳性例数分别为167、112、46;阳性率为96．O％、64．4％、26．4％,CA16：EV71为2．43：1。（2）首诊血清CA16和EV71IgM阳性例数为51、25,阳性率为38．9％、19．1％,复诊阳性例数为98、32,阳性率为74．8％、24．4％。（3）CA16VP1区核苷酸同源性为88．7％-98．5％,EV71VP1区核苷酸同源性为94．9％-99．7％,与c4亚型参比序列核苷酸同源性92．1％～95．3％。结论2009年夏季本院手足口病患儿病原以CA16、EV71为主,EV71阳性率较之前报道有较大幅度升高。EV71病毒株以c4亚型为主。实时RT-PCR法较血清学检测特异性IgM抗体更适于疾病早期诊断。     

深圳市某地2011年正常人群手足口病隐性感染调查研究

目的了解深圳市某地2011年成人及健康儿童手足口病隐性感染情况。探讨健康人群隐性感染与儿童患手足ISl病的关系。方法随机选取该区域幼儿同学生和老师共150人,分为0～、2～、5-岁组各30人,15～、24～、〉50岁组各20人。每人采集粪便标本5～8g,进行肠道病毒通用型、肠道病毒71型（EV71）、柯萨奇病毒A16型（CA16）核酸检测。结果共采集150份样本,EV通用型阳性13份（8．67％）,其中5岁以下的有11例（84．6％）。CA16阳性2份,EV71阳性1份,均属于0-岁组。结论肠道病毒在人群中隐性感染较为普遍,5岁以内儿童为主要感染人群。健康人群中引起手足口病常见的EV71和CA16阳性肠道病毒隐性感染率低。     

实时荧光PCR检测手足口病病原体方法的建立及初步应用

目的 建立一种能同时检测肠道病毒、肠道病毒71型和柯萨奇病毒16型等手足口病病原体核酸的实时荧光PCR方法.方法 针对上述病毒的基因序列,设计、合成引物、探针.对38例手足口病患者在实时荧光PCR仪上进行扩增、检测和结果分析;并与常规RT-PCR方法的检测结果进行比对.结果 38例患者肠道病毒、肠道病毒71型和柯萨奇病毒16型实时荧光PCR方法与常规RT-PCR方法的阳性率分别为:73.7%、60.5%、13.2%、71.1%、55.3%、13.2%.统计分析表明,两种方法差异不具有统计学意义.结论 成功建立了肠道病毒、肠道病毒71型和柯萨奇16型病毒核酸的实时荧光PCR检测方法.     

实时荧光PCR检测手足口病病原体方法的建立及初步应用

目的 建立一种能同时检测肠道病毒、肠道病毒71型和柯萨奇病毒16型等手足口病病原体核酸的实时荧光PCR方法.方法 针对上述病毒的基因序列,设计、合成引物、探针.对38例手足口病患者在实时荧光PCR仪上进行扩增、检测和结果分析;并与常规RT-PCR方法的检测结果进行比对.结果 38例患者肠道病毒、肠道病毒71型和柯萨奇病毒16型实时荧光PCR方法与常规RT-PCR方法的阳性率分别为:73.7%、60.5%、13.2%、71.1%、55.3%、13.2%.统计分析表明,两种方法差异不具有统计学意义.结论 成功建立了肠道病毒、肠道病毒71型和柯萨奇16型病毒核酸的实时荧光PCR检测方法.     

2009年徐州地区疑似肠道病毒感染病例病原学研究

目的 了解2009年徐州地区肠道病毒感染引起手足口病情况,为手足口病防控工作提供科学依据.方法 采用荧光RT-PCR方法对240例临床诊断为肠道病毒感染的手足口病例中的222例肛拭子和咽拭子标本同时进行EV和EV71、CA16特异性RNA检测,用ELA方法对其中的114例急性期血清标本进行EV71-IgM抗体检测.同时检测健康儿童254份咽拭子和258份血清标本以及密切接触者咽拭子54份标本.结果 240例疑似肠道病毒感染者,总EV感染率为72.50%(174/240),其中EV71感染率为57.92%(139/240),CoxA16感染率为9.17%(22/240),其他EV感染率为5.42%(13/240).240例疑似肠道病毒感染者中的222份肛拭子标本EV71-RNA阳性率为45.94%(102/222)、咽拭子EV71-RNA阳性率为25.68%(57/222),114份急性期血清中EV71-IgM抗体阳性率为86.84%(99/114).254份健康儿童咽拭子EV71-RNA阳性率为1.57%(4/254),未检出CoxA16-RNA;258份健康儿童血清标本EV71-IgM抗体阳性率是2.71%(7/258).密接(医护人员)咽拭子54份,EV-RNA检测均为阴性.结论 徐州地区手足口病流行主要由EV71型引起.3岁以下儿童流行期间存在被EV71型隐性感染,并产生IgM抗体.成人即使密切接触传染源也不易被感染EV71.疑似病例急性期血清中EV71-IgM抗体检测阳性率最高(86.84%),其次为肛拭子EV71(45.94%)和咽拭子EV71(25.68%)的RNA检测阳性率.ELA试剂盒用于EV71型感染的早期诊断,具有操作简单快速经济,值得在基层推广.     

实时荧光定量RT-PCR法定量检测手足口病病原体的探讨

目的了解北京市门头沟区手足口病患儿感染病毒的型别,探索用荧光定量RT—PCR法对手足口病患者咽拭子标本中肠道病毒71型（EV71）、柯萨奇病毒A16（CoxA16）型病毒载量进行定量检测的可行性。方法采用实时荧光RT—PCR体外扩增法对81例手足口病患儿咽拭子标本提取的RNA进行检测。结果28例手足口病患者体内CoxA16病毒载量〉10^3 copies·ml-1。实时荧光定量RT—PCR法构建的标准曲线显示,样本阈值环数（Ct）值与病毒拷贝数的对数（10g10）之间的相关系数为0．9998,相关性良好。4例手足口病患者咽拭子标本中EV71型病毒载量〉10^3 copies·ml-1。实时荧光定量RT-PCR法标准曲线显示,Ct值与病毒拷贝数的对数之间的相关系数为0．9996,相关性较好。结论门头沟区手足口病病原谱以CoxA16型为主,34．6％的手足口病患儿CoxA16型病毒载量〉10^3 copies·ml-1,4．9％的患儿EV71型〉10^3 copies·ml-1。实时荧光定量RT-PCR法对咽拭子标本中CoxA16、EV71核酸定量检测比较简便快速、结果直观、稳定性强,为进一步探讨患者体内病毒载量与临床症状的关系打下了良好的基础。     

2009年徐州地区疑似肠道病毒感染病例病原学研究

目的 了解2009年徐州地区肠道病毒感染引起手足口病情况,为手足口病防控工作提供科学依据.方法 采用荧光RT-PCR方法对240例临床诊断为肠道病毒感染的手足口病例中的222例肛拭子和咽拭子标本同时进行EV和EV71、CA16特异性RNA检测,用ELA方法对其中的114例急性期血清标本进行EV71-IgM抗体检测.同时检测健康儿童254份咽拭子和258份血清标本以及密切接触者咽拭子54份标本.结果 240例疑似肠道病毒感染者,总EV感染率为72.50%(174/240),其中EV71感染率为57.92%(139/240),CoxA16感染率为9.17%(22/240),其他EV感染率为5.42%(13/240).240例疑似肠道病毒感染者中的222份肛拭子标本EV71-RNA阳性率为45.94%(102/222)、咽拭子EV71-RNA阳性率为25.68%(57/222),114份急性期血清中EV71-IgM抗体阳性率为86.84%(99/114).254份健康儿童咽拭子EV71-RNA阳性率为1.57%(4/254),未检出CoxA16-RNA;258份健康儿童血清标本EV71-IgM抗体阳性率是2.71%(7/258).密接(医护人员)咽拭子54份,EV-RNA检测均为阴性.结论 徐州地区手足口病流行主要由EV71型引起.3岁以下儿童流行期间存在被EV71型隐性感染,并产生IgM抗体.成人即使密切接触传染源也不易被感染EV71.疑似病例急性期血清中EV71-IgM抗体检测阳性率最高(86.84%),其次为肛拭子EV71(45.94%)和咽拭子EV71(25.68%)的RNA检测阳性率.ELA试剂盒用于EV71型感染的早期诊断,具有操作简单快速经济,值得在基层推广.     

Schizophrenia in a North Swedish geographical isolate, 1900–1977. Epidemiology, genetics and biochemistry

Over 200 schizophrenic patients belonging to three major and interrelated pedigree complexes have been investigated over the past 30 years in a North Swedish geographically isolated population, presently numbering about 6,000. An intensive investigation of a number of biochemical correlates and genetic markers in a few selected families belonging to one of the major pedigrees has indicated new strategies for the current research program.
Schizophrenia, as defined operationally, is significantly associated with decreased activities of two enzymes (1) blood platelet monoamine oxidase, (2) plasma dopamine-β-hydroxylase, and (3) with the genetic marker Gc² (group specific antigen). Both enzymes are subject to genetic variation. A positive score for linkage between schizophrenia and low plasma DBH activity has been calculated, but, so far, available data are insufficient for discrimination between linkage and partial contribution of genetically controlled low plasma DBH to the pathogenesis of the disease. Alternatively, both mechanisms could be involved.
As a model for continued research, schizophrenia is explained as based on a double dominant-recessive genotype (Aabb), representing a vulnerability which in about 50 % of cases develops into clinical schizophrenia. It is suggested that the dominant mutation (A) operates on or affects MAO activity, and that the recessive genotype (bb) is instrumental in low variates of DBH activity and very likely such variates within the normal range of physiological variation. Moreover, it is suggested that the combined effects of MAO- and DBH-reduced efficiency on the metabolism of e.g. dopamine could be an essential pathogenic mechanism for the schizophrenic illness which is segregating in this population.     

Renal dysplasia and asplenia in two sibs

A family is reported in which two sibs, one male and the other female, both died within 24 hours of birth with enlarged polycystic kidneys. Postmortem histology in the second child showed gross renal dysplasia. In both children the pancreas was enlarged, nodular and cystic but the liver appeared macroscopically normal. In the second child, histological examination confirmed pancreatic fibrosis with cystic dilation of ducts, but showed portal fibrosis with bile duct proliferation in the liver.
This combination of findings is very reminiscent of those in a girl and her brother reported by Ivemark et al. (1959). The children reported here also showed absence or hypoplasia of the spleen, cardiac anomalies and other features of the Ivemark syndrome (Ivemark 1955), a quite different, usually sporadic, congenital disorder. It is suggested that the children described here have a distinct lethal congenital disorder, probably inherited in an autosomal recessive manner.     

The dominant diseases of modernized societies as omega-3 essential fatty acid deficiency syndrome: Substrate beriberi

About 1900, modern food selection and processing caused widespread $\text{epidemics}$ of the B vitamin deficiency diseases of beriberi and pellagra which, for genetic reasons, often expressed as different diseases ranging from bowel and heart disease to dermatoses and psychoses. But the B vitamins merely help convert essential fatty acids (EFA) into the prostaglandin (PG) tissue regulators and it now turns out that, through hydrogenation, milling and selection of w3-poor southern foods, we have also been systematically depleting, by as much as 90%, a newly discovered trace Nordic EFA (w3) of special importance to primates and sole precursor of the PG3(4) series, even as a concurrent fiber deficiency increases body demand for EFA. Since substrate EFA is processed by many B vitamin catalysts, an EFA deficiency will mimic a $\text{panh}$ypovitaminosis B, i.e., a mixture of $\text{substrate}$ beriberi and $\text{substrate}$ pellagra resembling vitamin beriberi and pellagra but exhibiting as even more diverse $\text{endemic}$ disease. This would consitute a second stage of the Modern Malnutrition and explain why some workers now hold the dominant diseases of modermized societies to be new, nutritionally based, pellagraform yet lipid-related and to range, once again, from heart disease to psychosis. It is an assumption that our dominant diseases are unrelated to each other or are merely revealed by our diagnostic acumen and therapeutic success; and that hydrogenating millions of tons of food oils annually, to destroy the rancidity producing w3-EFA, is safe for $\text{primates}$. Extensive beriberiform disease is reported here in 32 typical cases taken from medical practice which responds strikingly to linseed oil supplements (60% w3-EFA) in confirmation of identical results in Capuchins.     

'Very sore nights and days': the child's experience of illness in early modern England, c.1580-1720

Sick children were ubiquitous in early modern England, and yet they have received very little attention from historians. Taking the elusive perspective of the child, this article explores the physical, emotional, and spiritual experience of illness in England between approximately 1580 and 1720. What was it like being ill and suffering pain? How did the young respond emotionally to the anticipation of death? It is argued that children’s experiences were characterised by profound ambivalence: illness could be terrifying and distressing, but also a source of emotional and spiritual fulfilment and joy. This interpretation challenges the common assumption amongst medical historians that the experiences of early modern patients were utterly miserable. It also sheds light on children’s emotional feelings for their parents, a subject often overlooked in the historiography of childhood. The primary sources used in this article include diaries, autobiographies, letters, the biographies of pious children, printed possession cases, doctors’ casebooks, and theological treatises concerning the afterlife.     

Guidelines for the Validation and Use of Immunoassays for Determination of Introduced Proteins in Biotechnology Enhanced Crops and Derived Food Ingredients

Recent advancements in agricultural biotechnology have created a need for analytical techniques to determine introduced proteins in crops enhanced through modern biotechnology techniques. These proteins are expressed in plant tissues and may be present in food ingredients. Immunoassays are ideally suited for protein detection and may be used as both quantitative and threshold methods. Microplate ELISA and lateral flow devices are two of the most commonly used immunoassay formats for agricultural biotechnology applications. This paper provides general background information and a discussion of criteria for the validation and application of immunochemical methods to the analysis of proteins introduced into plants and food ingredients using biotechnology methods. It is the result of a collaborative effort of members of the Analytical Environmental Immunochemical Consortium. This collaborative effort represents the combined expertise of several organizations to reach consensus on establishing guidelines for the validation and use of immunoassays. Further, the paper offers developers and users a consistent approach to adopting the technology as well as aid in producing accurate and meaningful results.     

Ultracryotomy: development and application (with examples from the yeast cytology) (author's transl)

The preparation steps usually necessary for obtaining ultrathin frozen sections of biological material (chemical prefixation, enclosing, cryoprotective treatment, freezing, sectioning, and post-staining the sections for transmission electron microscopy) are submitted to a critical analysis. The application of cryo-ultramicrotomy, in particularly for cytochemical purposes, is reviewed. Fundamental considerations of chemical prefixation and poststaining are supported by examples from yeast cytology. Furthermore, the efficiency of the cryo-ultramicrotomy (electron optical resolution of ultrastructural details) is demonstrated on yeast cells and protoplasts.     

HLA in North Colombia Chimila Amerindians

HLA-A,-B,-C,-DRB1 and -DQB1 alleles have been studied in Chimila Amerindians from Sabana de San Angel (North Colombian Coast) by using high resolution molecular typing. A frequent extended haplotype was found:HLA-A*24:02-B*51:10-C*15:02-BRB1*04:07-DQB1*03:02 (28.7%) which has also been described in Amerinndian Mayos Mexican population (Mexico, California Gulf, Pacific Ocean). Other haplotypes had already been found in Amerindians from Mexico (Pacific and Atlantic Coast), Peru (highlands and Amazon Basin), Bolivia and North USA. A geographic pattern according to HLA allele or haplotype frequencies is lacking in Amerindians, as already known. Also, five new extended haplotypes were found in Chimila Amerindians. Their HLA-A*24:02 high frequencies characteristic is shared with aboriginal populations of Taiwan; also, HLA-C*01:02 high frequencies are found in New Zealand Maoris, New Caledonians and Kimberly Aborigines from Australia. Finally, this study may show a model of evolutionary factors acting and rising one HLA allele frequency (-A*24:02), but not in others that belong to the same or different HLA loci.     

The universal muscular chamber. A basic unit of the genitourinary, cardiovascular, gastro-intestinal, skeletal, cutaneous, respiratory and other systems, endocameral hypertension; and spasm, the key to their idiopathic diseases and arthropathies

Starting with the integument, we see many organs are contractile sacs or multiples thereof, which tubes or bags constitute the major part of the entire body. Recognition of this basic unit and its characteristics sheds new light, individually and collectively, on many disorders previously considered unrelated. Muscular tears and perforations develop in the walls of these chambers, being no way peculiar to those organs, wherein, hydrochloric acid occurs. So, it is not necessary to explain the absence of excessive acid from patients who exhibit holes in the gastric, uterine, aortic, duodenal, rectal, pulmonary, retina, and other walls. Muscle, not acid is the great common factor relating idiopathic disorders in the gastrointestinal tract to each other and to similar diseases in other systems. When the units are linked together, the lesions tend to appear as arthropathies, i.e. at the joints. Rephrasing common-place observations, frees us from conventional, conceptual cul-de-sacs. An observation is only as good as its interpretation, so all possibilities must be considered, otherwise, we will remain blinded by our misconceptions.     

Der Einfluß von Nahrungsmitteln und Rauchen auf die klinisch-chemische Diagnostik von Phäochromozytom,Neuroblastom und Karzinoid-Syndrom

Zusammenfassung Der Einfluß von verschiedenen Nahrungsmitteln auf Methoden zur Bestimmung von Adrenalin (AD), Noradrenalin (NA), Vanillinmandelsäure (VMS), Metanephrinen (MN), Homovanillinsäure (HVS) und 5-Hydroxyindolessigsäure (5-HIE) im 24 h-Harn zur Diagnose des Phäochromozytoms bzw. Karzinoid-Syndroms wurde untersucht. Die in die Untersuchung einbezogenen Nahrungsmittel waren: Tee, Kaffee, Mandeln, Ananas, Käse, Walnüsse, Vanillepudding, Bananen, Tomaten und Milchschokolade. Außerdem wurde der Einfluß des Zigarettenrauchens auf die Bestimmung von AD, NA, VMS und MN untersucht.Walnüsse führten zu einer starken Erhöhung der 5-HIE-Ausscheidung. Bananen erhöhten die Ausscheidung von AD, NA, VMS, MN und 5-HIE. Kaffee und Ananas bewirkten eine geringe Zunahme der MN-Werte. Rauchen von 20–30 Zigaretten/Tag beeinflußte keine der vier Variablen.Wenn die beschriebenen Methoden benutzt werden, sollte lediglich auf den Verzehr von Bananen und Walnüssen vor und während der Harnsammelperioden verzichtet werden, da die oberen Normgrenzen im Harn überschritten werden könnten. Ein Verzicht auf Kaffee und Ananas in normalen Mengen ist nicht erforderlich. Es besteht kein Anlaß, weiterhin die bisherigen umfangreichen Restriktionen der übrigen Nahrungsmittel beizubehalten.