鲍曼不动杆菌临床株多重药物外排泵AdeABC的研究

目的 研究鲍曼不动杆菌(Acinetobacter baumannii)临床株外排泵AdeABC的表达与耐药的关系及表达调控.方法 微量肉汤稀释法检测鲍曼不动杆菌临床株对抗菌药物的敏感性及泵抑制剂作用,RT-PCR检测泵基因adeB的mRNA表达水平,PCR扩增泵调控基因adeRS并测序分析.结果 30株多重耐药的鲍曼不动杆菌临床株及5株敏感株均存在泵结构基因片段adeB和调控基因adeRS,随机选取15株多重耐药株中均检测到adeB的mRNA表达,而在5株敏感株中无表达.测定2株多重耐药株调控基因adeRS序列均出现基因突变,发生氨基酸替代及缺失.结论 鲍曼不动杆菌临床株外排泵AdeABC的表达可能与耐药性有关,在多重耐药株中存在着调控基因adeRS的基因序列变化.     

鲍曼不动杆菌多重耐药性与主动外排机制的相关性研究

目的 了解多重耐药鲍曼不动杆菌主动外排系统及双组分调节系统编码基因的携带情况,并观察外排泵抑制剂对多重耐药鲍曼不动杆菌耐药水平的影响程度,以探讨鲍曼不动杆菌多重耐药性与胞膜主动外排系统的关系.方法 PCR方法 扩增外排泵编码基因adeB及双组分调节系统编码基因adeR和adeS.采用琼脂稀释法测定50株多重耐药鲍曼不动杆菌对环丙沙星、阿米卡星、头孢噻肟和亚胺培南的最低抑菌浓度(MIC),并观察在含25μg/ml利血平条件下MIC值的变化程度.结果 50株多重耐药鲍曼不动杆菌adeB、adeR及adeS基因的携带率分别为94%、96%及92%.以环丙沙星、头孢噻肟、阿米卡星和亚胺培南作为底物,分别有49、50、50和46株菌在含25μg/ml利血平的条件下MIC值降低4倍或4倍以上,呈现明显的外排作用.结论 主动外排机制是本地区鲍曼不动杆菌多重耐药的重要原因之一.     

泛耐药鲍氏不动杆菌氨基糖苷类耐药相关基因研究

目的 探讨泛耐药鲍氏不动杆菌氨基糖苷类抗菌药物耐药机制。方法 收集2014年湖北省黄石市中心医院住院患者中分离的20株泛耐药鲍氏不动杆菌,用mdfA测序确认菌种,再采用聚合酶链反应法分析16种氨基糖苷类药物获得性耐药相关基因。 结果 20株泛耐药鲍氏不动杆菌共检出4种氨基糖苷类获得性耐药基因,氨基糖苷类修饰酶基因aac(3)-Ⅰ阳性率15.00％;ant(3")-Ⅰ阳性率20.00％;aph3'-Ⅰ阳性率100.00％;16SrRNA甲基化酶未检出;外排泵adeB基因阳性率95.00％。 结论　泛耐药鲍氏不动杆菌中携带的耐药基因和耐药表型相对应,对氨基糖苷类药物耐药与产aac(3)-Ⅰ、ant(3")-Ⅰ、aph3'-Ⅰ修饰酶基因和获得外排泵adeB 2种耐药机制有关。     

体内诱导耐碳青霉烯类鲍曼不动杆菌膜蛋白质组学研究

目的 研究膜蛋白在耐碳青霉烯类抗生素鲍曼不动杆菌中的作用.方法 从同一住院病人体内分别收集碳青霉烯类敏感和耐药的鲍曼不动杆菌各1株.经多序列位点测序分型(MIST)和细菌基因组外回文结构重复序列分型(REP-PCR)分析后,等电聚焦电泳检测其已知碳青霉烯类水解酶的表达,在此进行膜蛋白二维电泳和质谱鉴定分析,并最后应用PABN(Phe-Arg-β-naphthylamide)外排泵抑制剂检测其外排泵相关膜蛋白表达.结果 MIST和REP-PCR结果表明耐药株来源与敏感菌株属于同一型别;等电聚焦电泳在P17.6和P19.0处两株菌中检测到β-内酰胺酶,没有检测到任何已知的碳青霉烯类水解酶;耐约株和敏感株的差异膜蛋白组学鉴定出相对分子质量(M_r)为34×10~3外排泵膜蛋白和0prD与CarO膜孔蛋白,且后续的外排泵抑制剂试验表明,在PAβN存在的情况下,耐药株的亚胺培南最低抑菌浓度(MIC)由大于32 μg/ml下降到8 μg,/ml.结论 本研究发现外排泵膜蛋白的过度表达伴随OprD和CarO膜孔蛋白的下调是临床分离耐碳青霉烯类抗生素鲍曼不动杆菌主要耐药机制.     

医院泛耐药鲍曼不动杆菌对碳青霉烯抗生素的耐药机制

目的 探讨医院泛耐药鲍曼不动杆菌对碳青霉烯抗生素的耐药机制.方法 应用PCR方法对2010年12月至2012年3月期间本院从临床痰标本中分离的36株泛耐药鲍曼不动杆菌进行碳青霉烯酶IMP、OXA23基因和整合子基因检测;提取细菌膜蛋白行SDS-PAGE电泳分析其组成.结果 36株泛耐药鲍曼不动杆菌碳青霉烯酶OXA23基因扩增均为阳性;14株碳青霉烯酶IMP基因扩增阳性,22株阴性.12株整合子PCR产物约1200 bp,10株约3000 bp,14株整合子PCR产物约3500 bp.与碳青霉烯抗生素敏感鲍曼不动杆菌膜蛋白比较,22株泛耐药鲍曼不动杆菌存在相对分子质量为25 000、36 000的膜蛋白缺失.结论 医院泛耐药鲍曼不动杆菌耐碳青霉烯抗生素机制与产IMP、OXA23碳青霉烯酶及膜蛋白缺失有关.     

60株鲍曼不动杆菌耐药基因携带情况分析

目的研究临床分离的60株鲍曼不动杆菌耐药谱及Ⅰ型整合子和β-内酰胺酶等基因携带情况。方法用微量肉汤稀释法测定16种临床常用抗菌药物的最小抑菌浓度;PCR检测β-内酰胺酶、Ⅰ型整合子和外排泵基因,对阳性基因进行序列分析。结果60株菌中,多重耐药株53株,占88.3%;6株携带OXA-23基因,均对包括碳青霉烯在内的5类以上抗菌药耐药,并具有高耐药特性;38株携带PER-1基因,对头孢菌素类耐药率显著高于PER-1基因阴性菌株(P<0.01);45株检出Ⅰ型整合子结构基因,多重耐药率明显高于Ⅰ型整合子阴性菌株(P<0.01);Ⅰ型整合子和PER-1基因同时阳性25株,与7株两者同为阴性菌株相比,多重耐药率增高(P<0.01),但耐药程度无显著差别。结论Ⅰ型整合子基因及β-内酰胺酶类基因的作用是导致鲍曼不动杆菌多重耐药的重要原因;OXA-23基因阳性菌株多为泛耐药和高耐药株,有必要采取有效措施控制其传播。     

2014～2015年鲍曼不动杆菌的医院分离情况和耐药性分析

目的 了解2014年至2015年鲍曼不动杆菌在本院的分布情况,并对其耐药性进行分析,为临床合理使用抗菌药物及控制院内感染提供依据.方法 收集本院2014年1月至2015年12月间送检的各种临床标本,应用VITEK 2 Compact 全自动微生物分析系统对临床分离的病原菌进行鉴定,同时用K-B药敏纸片法测定抗菌药物敏感率,结果按照CLSI标准判定.利用WHONT 5.6软件分析鲍曼不动杆菌的耐药性及其分布.结果 重症监护病房(ICU)和呼吸科的鲍曼不动杆菌分离率最高,且以呼吸道标本为主,达90％以上;2014至2015年我院共分离出鲍曼不动杆菌212株,耐药结果显示,鲍曼不动杆菌对氨曲南、头孢他啶耐药明显,超过40％,对头孢呋辛钠表现出100％耐药,但亚胺培南、头孢哌酮-舒巴坦、复方磺胺甲恶唑对鲍曼氏不动杆菌仍有较好的抗菌作用(耐药率＜25％),其中鲍曼不动杆菌对头孢哌酮-舒巴坦的耐药率最低,耐药率小于5％.结论 鲍曼不动杆菌感染率不断上升,已成为ICU感染的主要病原菌,但由于其耐药性各不相同,对其进行耐药性分析,可以进一步指导临床合理使用抗生素,控制多重耐药鲍曼不动杆菌传播.     

鲍曼不动杆菌临床分离株表型、基因型和耐药基因的对比研究

目的 对比研究鲍曼不动杆菌临床分离株基因型和编码耐药基因的差异,并分析其与临床多重耐药性的关系.方法 随机收集中南大学湘雅二医院2008年9月至2009年9月分离的77株鲍曼不动杆菌,采用WHO推荐的K-B法对鲍曼不动杆菌进行临床常见15种抗生素药物敏感试验,并对药敏谱进行分析.用随机扩增多态性DNA法(RAPD)技术进行基因分型.并利用PCR对β-内酰胺酶基因TEM-1、IMP、OXA-23、OXA-24、AmpC和氨基糖苷类修饰酶基因aac(3)-Ⅰ、aac(6')-Ⅰ、ant(3")-Ⅰ和16S rRNA甲基化基因armA、rmtA、rmtB进行扩增及序列分析.对比分析鲍曼不动杆菌耐药基因的携带情况,以及与基因型和耐药性的关系.结果 77株鲍曼不动杆菌中敏感菌株有31株,对5种或5种以上抗生素耐药的多重耐药菌株46株,内含全耐药菌株10株.RAPD技术将其分为17型,为A-G型,多重耐药株中E型为优势克隆株(17株),在重症监护病房(ICU)中流行最广,占47.1%(8/17).敏感株中各型散在分布.PCR扩增结果显示,多重耐药株和敏感株携带TEM-1、IMP、OXA-23、OXA-24、AmpC、aac(3)-Ⅰ、aac(6')-Ⅰ、ant(3")-Ⅰ和armA耐药基因的比率分别为95.7%、39.1%、84.8%、54.3%、87.0%、89.1%、84.8%、45.7%、63.0%和58.1%、9.7%、32.3%、48.4%、48.4%、29.0%、45.2%、12.9%、9.7%,未发现rmtA和rmtB基因阳性菌株.经x2检验,除OXA-24外,其余各耐药基因携带率比较差异有统计学意义(P＜0.05).药敏分析提示携带以上耐药基因的鲍曼不动杆菌菌株的耐药率明显高于未携带该基因的菌株,其中对阿米卡星和庆大霉素耐药的菌株,其氨基糖苷类酶基因均阳性(34.8%),含所测的所有β-内酰胺酶基因的菌株均为全耐药株.结论 与临床分离的敏感鲍曼不动杆菌相比,多重耐药株耐药谱广,耐药率高,其携带β-内酰胺酶基因和氨基糖苷类酶基因种类多,分离率高,且同一克隆的多重耐药株可在病室内和病室间传播.     

鲍曼不动杆菌armA基因的分布与耐药性的研究

目的 调查我院鲍曼不动杆菌中16S rRNA甲基化基因armA的分布以及与鲍曼不动杆菌耐药谱的关系,并初步探讨其在分子流行病学分析中的作用.方法 收集72株鲍曼不动杆菌,采用K-B法对鲍曼不动杆菌进行药物敏感试验,后采用PCR筛选鲍曼不动杆菌的16S rRNA甲基化基因armA,并利用随机扩增多态性DNA法(RAPD)技术进行基因分型.统计各鲍曼不动杆菌菌株对多种氨基糖苷类药物的药敏结果,并分析基因型与耐药性的关系.结果 根据PCR产物片段大小,72株鲍曼不动杆菌共有armA基因阳性菌株20株(27.8%).含有armA基因型鲍曼不动杆菌菌株对庆大霉素、妥布霉素和阿米卡星的耐药率均为90%;随机扩增多态性DNA法显示20株armA基因阳性的鲍曼不动杆菌主要分为7型,A型为优势克隆株.结论 产16S rRNA甲基化基因armA的鲍曼不动杆菌菌株可对多种氨基糖苷类抗生素高水平耐药.同一克隆菌株在病房内和病房间的传播为我院armA基因传播的主要方式.     

亚胺培南耐药鲍曼不动杆菌16S rRNA甲基化酶基因检测及流行菌株分析

目的 研究我国多中心亚胺培南耐药的鲍曼不动杆菌的耐药性、16S rRNA甲基化酶的阳性率及分子流行病学特征.方法 收集2004年11月-2005年11月国内6省市19家医院临床分离的342株亚胺培南耐药的鲍曼不动杆菌.采用琼脂稀释法和E test法对18种抗菌药物测定分离株的最低抑菌浓度(MIC)值;脉冲场凝胶电泳(PFGE)分析菌株的同源性;PCR法筛选4种16S rRNA甲基化酶基因armA、rmtA、rmtB、rmtC,克隆测序明确基因型;接合试验、质粒抽提、电转化以及Southern blot确定甲基化酶耐药基因定位.结果 所有菌株均为多重耐药株,其中对阿米卡星、庆大霉素、妥布霉素、异帕米星、奈替米星耐药率分别为92.6%、98.6%、87.4%、90.9%和92.4%.PCR扩增、测序证实221株鲍曼不动杆菌检出甲基化酶armA基因;另3种甲基化酶基因rmtA、rmtB、rmtC均阴性.在上述342株鲍曼不动杆菌临床分离株中,298株可归类为6个流行克隆,44株为散发株.3个主要克隆(A、B、C)在全国广泛播散,分别在国内6家、3家、11家医院内流行.接合试验、质粒抽提、电转化及Southern blot表明armA编码基因存在于染色体上.结论 亚胺培南耐药鲍曼不动杆菌的表型均为多重耐药.介导氨基糖苷类抗生素高度耐药的16S rRNA甲基化酶基因armA在鲍曼不动杆菌中广泛存在,其主要传播方式为克隆播散,这必将引起临床的高度关注.     

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.     

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.     

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.     

What will studies of Fulani individuals naturally exposed to malaria teach us about protective immunity to malaria?

There are an estimated over 200 million yearly cases of malaria worldwide. Despite concerted international effort to combat the disease, it still causes approximately half a million deaths every year, the majority of which are young children with Plasmodium falciparum infection in sub-Saharan Africa. Successes are largely attributed to malaria prevention strategies, such as insecticide-treated mosquito nets and indoor spraying, as well as improved access to existing treatments. One important hurdle to new approaches for the treatment and prevention of malaria is our limited understanding of the biology of Plasmodium infection and its complex interaction with the immune system of its human host. Therefore, the elimination of malaria in Africa not only relies on existing tools to reduce malaria burden, but also requires fundamental research to develop innovative approaches. Here, we summarize our discoveries from investigations of ethnic groups of West Africa who have different susceptibility to malaria.     

'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.     

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.     

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.     

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.