MRSA菌20株耐药性及耐药基因研究

目的对MRSA菌耐药性及耐药基因进行研究。方法对20株MRSA菌的耐药性和β内酰胺类耐药相关基因(mecA、TEM)、氨基糖甙类耐药相关基因（aac(6’)／aph(2”)、aph(3’)-Ⅲ、ant(3”)-Ⅰ、ant(2”)-Ⅰ、ant(6)-Ⅰ）、四环素耐药相关基因(tetM)等主要耐药基因进行检测。结果20株MRSA药敏试验除万古霉素、替考拉宁敏感外，青霉素、苯唑西林、头孢唑林、头孢呋辛、亚胺培南、庆大霉素、四环素、红霉素、环丙沙星等均耐药。20株MRSA菌均检出mecA、TEM、aac(6’)／aph(2”)、tetM等β内酰胺类、氨基糖甙类、四环素类、红霉素类耐药相关基因，其中6株还同时检出aph(3’)-Ⅲ基因。结论全部菌株均已携带β内酰胺类、氨基糖甙类、四环素类耐药相关基因。     

地中海诺卡菌的菌丝形态特征与产生利福霉素B的定量分析

在各种放线菌中,抗生素产生的起动(生理分化)与菌丝形态变化(形态分化)有密切关系。然而,高度分支的菌丝会增加培养基的粘度,影响正常的通气和混合,因而成为优化抗生素生产工艺中的主要问题。 利福霉素B系地中海诺卡菌产生。这种菌株的形态特征明显受培养条件的影响,而利福霉素B的产生又同形态变化密切相关。Lee等在分批培养中利用不同的生产培养基     

一株多重耐药铜绿假单胞菌耐药基因的研究

目的:研究铜绿假单胞菌多重耐药情况和相关机制。方法:采用聚合酶链式反应(PCR)法对一株多重耐药铜绿假单胞菌进行β-内酰胺酶基因、氨基糖苷类修饰酶基因、氯霉素类基因、耐消毒剂和磺胺类基因检测。结果:PCR结果显示该株PA的TEM,MIR,CTX-M1群,OXA-1群,OXA-10群,aac(6′)-b,aac(6′)-,aac(3)-,ant(2″)-,catB,qacE△1-sul1基因均为阳性,GES,DHA,VEB,cmlA基因均阴性,Opr D2未缺失。结论:该株铜绿假单胞菌存在多重耐药基因,而且对胺类、胍类消毒剂耐受。     

铜绿假单胞菌“泛耐株”耐药相关基因研究

目的 研究"泛耐"铜绿假单胞菌中多种耐药基因的存在情况及耐药机制.方法 采用聚合酶链反应(PCR)法检测22株"泛耐"铜绿假单胞菌的β-内酰胺酶基因、氨基糖苷类修饰酶(AMEs)基因、喹诺酮类耐药基因、耐消毒剂基因(qacE△l-sull)和整合酶基因.结果 PCR扩增结果显示22株菌中oprD2、blaCAEB、gyrA、aac(6')-Ⅰb、ant(2")-Ⅰ和qacE △ J-sull基因均阳性;blaSHV、blaSPM、blaGIM、blaDHA、blaOXA-10、blaGES、blaPER、aac(3')-Ⅰ和aac(6')-Ⅱ基因均阴性;blaIMP、blaTEM、blaVEB、aac(3')-Ⅱ、ant(3")-Ⅰ、int Ⅰ1和blaVIM阳性率分别为95.5%、77.3%、13.6%、4.5%、4.5%、4.5%和4.5%.结论 oprD2基因阳性说明这些泛耐菌株中外膜蛋白并未缺失,多种耐药基因同时存在是本研究中铜绿假单胞菌"泛耐"的根本原因.     

致病性猪沙门菌四环素耐药基因tetB的PCR检测

目的检测临床分离猪源致病性沙门菌四环素的耐药基因,确定沙门菌四环素耐药基因类型。方法用KB法测定分离株对四环素等21种抗生素的耐药情况,用PCR方法检测四环素耐药基因tetA、tetB、tetC、tetD、tetE、tetG及tetK,并对扩增产物进行测序。结果临床分离株全部对四环素耐药,33株沙门菌中有29株扩增出tetB特异性片段,基因序列与参考菌的同源性为99%,tetB检测与药敏试验阳性符合率为87.9%。结论tetB的PCR检测对四环素耐药性具有较高的特异性,tetB基因是决定本试验中临床分离株四环素耐药的主要基因,为四环素耐药性的分子流行病学监测提供了依据。     

阴道加德纳菌四环素耐药基因tetM的体外扩增及其产物的序列测定

利用聚合酶工反应（ＰＣＲ）对５株耐四环素阴道加德纳菌（Ｇａｒｄｎｅｒｅｌｌａｖａｇｉｎａｌｉｓ（ＭＩＣ〉１６ｍｇ／Ｌ）的耐药基因ｔｅｔＭ进行体外扩增。核苷酸序列测定证实，１株耐四环素阴道加德纳功的ｔｅｔＭ核苷酸序列与另４株ｔｅｔＭ的序列不同，两者的同源性为９８．４％，同时又和Ｔｎ１５４５、Ｔｎ９１６部分同源。说明在阴道加德纳菌中ｔｅｔＭ基因存在多态性，其获得方式，来源途径均不相同。     

多重PCR技术检测沙门菌两种四环素耐药基因

应用两对特异性引物同时检测四环素耐药基因tetB和tetC通过对35株沙门菌分离株的四环素耐药性检测，表明所有沙门菌分离株均含tetC基因，与药敏试验结果阳性符合率65．7％，8株同时含有tetB基因，与药敏试验结果阳性符合率100％，tetB基因和tetC基因双阳性的菌株与药敏试验结果阳性符合率也为100％。取其中部分菌株扩增出tetB和tetC基因片段进行序列分析，5株菌的tetB基因扩增产物序列完全相同，与质粒pRT11相应序列同源性达99．7％；14株菌的tetC基因扩增产物与质粒pBR322中的相应序列同源性为100％。证实沙门菌耐药基因普遍存在，且同时含有tetB和tetC基因的菌株表现耐药。多重PCR技术同时检测两种四环素耐药基因，适合大量样本的检测，对开展沙门菌四环素多种耐药基因的分子流行病学监测提供了有效途径。     

铜绿假单胞菌对环丙沙星耐药与喹诺酮类耐药基因的关系研究

目的:探讨喹诺酮耐药(QNR)基因介导的铜绿假单胞菌对环丙沙星的耐药性。方法:用K-B纸片法测定30株铜绿假单胞菌对环丙沙星、庆大霉素、哌拉西林、头孢噻肟4种抗菌药物的敏感性。PCR法检测(QNR)基因。结果:铜绿假单胞菌对环丙沙星的耐药率为26.70%,1株环丙沙星耐药性铜绿假单胞菌携带有QNR基因。结论:铜绿假单胞菌对环丙沙星的耐药性与QNR基因有密切相关性。     

冷泉来源诺卡氏菌OUCLQ19-35-1次级代谢产物的研究

目的 探究深海冷泉来源微生物的次级代谢产物产生能力,从中挖掘具有抗多重耐药(multi-drug resistant, MDR)菌活性的次级代谢产物,为新药研发提供化合物实体。方法 采用稀释涂布法分离纯化深海冷泉海泥样品中的放线菌,通过琼脂扩散法筛选具有抗MDR菌活性的放线菌;基于16S rRNA基因片段序列分析和系统进化树构建初步确定目标放线菌种属;对目标放线菌进行大规模发酵,采用有机溶剂萃取、反相硅胶柱层析、半制备高效液相等分离手段对发酵产物进行分离纯化,利用NMR、MS等波谱学技术并结合文献对化合物进行结构鉴定,然后对化合物进行抗MDR菌活性测试。结果 从深海冷泉中筛选到一株具有抗MDR菌Micrococcus luteus ML01和Staphylococcus aureus CCARM3090活性的放线菌OUCLQ19-35-1,16S rRNA序列及系统进化树分析初步确定其为Nocardiopsis synnemataformans;从其发酵产物中分离得到3个化合物,分别为questiomycin A(1)、1,6-dihydroxyphenazine(2)和5a,6,11a,12-tetrahydro-5a,11a-dimethyl[1,4]benzoxazino[3,2-b][1,4]benzoxazine(3);活性结果显示,化合物1-3均无抗MDR菌活性,但其所在的组分有抑菌活性。结论 从深海冷泉筛选得到一株诺卡氏菌OUCLQ19-35-1,其能够产生抗MDR菌的活性次级代谢产物,具有潜在的应用价值,但其活性成分待进一步的确定。     

碳青霉烯类耐药肺炎克雷伯菌临床分离株中质粒介导的喹诺酮类耐药基因检测

目的：探讨质粒介导的喹诺酮类耐药（PMQR）基因在碳青霉烯类耐药肺炎克雷伯菌（CRKP）临床分离株中的分布情况。方法：收集CRKP29株,聚合酶链反应（PCR）扩增并确定产物基因型。结果：CRKP中PMQR基因检出率为48．3％（14／29）,其中qnrB5株,包括qnrB21株、qnrB43株、qnrB101株;qnrS 5株,均为qnrS1;1株同时携带qnrS1和qnrB4;有3株携带aac（6′）-Ib-cr基因。所有PMQR基因阳性菌株均同时携带β-内酰胺酶基因,其中8株同时携带碳青霉烯酶基因,占57．1％（8／14）,以blaKPC-2（4／14）及blaNDM-1（3／14）为主。结论：PMQR基因在临床分离的CRKP中较为普遍,以qnr基因为主,且qnr阳性菌株碳青霉烯酶基因携带率较高,均为多重耐药株。     

Genetic analysis of Tn916-like elements conferring tetracycline resistance in clinical isolates of Clostridium difficile

As an important clinically relevant pathogen, Clostridium difficile has a high multidrug resistance rate. Conjugative transposons play a vital role in its resistance phenotype. In the present study, 34 tetracycline-resistant clinical isolates of C. difficile were studied to detect tetracycline resistance genes and the presence of transposons. Thirty-two isolates were found to harbour Tn916-like elements carrying the tet(M) resistance gene, of which only one copy existed in the genome by Southern blot analysis. To characterise the genetic organisation of the Tn916-like elements, overlap PCR assays were performed with nine primer pairs, revealing three types of elements designated T1 to T3. The prevalent element T1 lacking PCRA (ORF₂₃ to ORF₂₁) and PCRB (ORF₂₁ to ORF₂₀) products, present in the epidemic ST37 clone, was further analysed by genome walking PCR in the left and right end sequences of the novel Tn916-like element. A gene coding for an FtsK/SpoIIIE family protein was found to replace the ORF24 to ORF21 region in Tn916. Moreover, the element could hardly conjugate between cells by filter mating experiments. These findings suggest that the dissemination of Tn916-like elements in epidemic ST37 strains in China was likely to have been conferred by clonal spread, signifying the importance of future surveillance and characterisation of conjugative transposons.     

耐亚胺培南铜绿假单胞菌的耐药及基因分析

目的分析耐亚胺培南铜绿假单胞菌（IRPA）的耐药性及耐药基因。方法聚合酶链反应（PCR）法对IRPA耐药相关基因（IMP、SPM、VIM、GIM、GES、OXA-10）和膜微孔蛋白基因OprD2进行检测;用脉冲场凝胶电泳进行菌株同源性检测;用VITEK2Compact进行细菌鉴定及药敏试验。结果28株IRPA对哌拉西彬他唑巴坦、妥布霉素耐药率均为32．1％,对环丙沙星、左旋氧氟沙星耐药率分别为46．4％和67．9％,对其他抗菌药物均呈较高的耐药性（82．1％～100．0％）;有4株对16种抗菌药物均耐药。OprD2基因为25．0％（7／28）,SPM基因为14．3％（4／28）,OXA-10基因为3．6％（1／28）;VIM、GIM、GES耐药基因未检出。结论IRPA的多重耐药现象较严重,治疗IRPA引起的感染首选哌拉西林／他唑巴坦、妥布霉素;耐药基因以IMP、OprD2为主。     

血压盐敏感性及其易感基因

盐敏感（SS）是人群中个体间对限盐措施或盐负荷呈现血压升高或降低的反应,是个体对高血压的遗传易感性,也是影响高血压及其并发症发生发展的一个重要因素。盐敏感性的早期识别诊断、盐敏感性易感基因及其遗传因素等的研究对高血压发病机制及防治措施的研究有重要意义。目前关于血压盐敏感性易感基因的研究报道中存在较大差异,可能与种族差异、饮食习惯、环境因素、基因环境交互作用及样本含量等有关。本文对近年国内外关于盐敏感性易感基因的研究进行综述,可作为血压盐敏感性研究的候选基因有钠泵基因、α-adducin基因、胰岛素受体13亚单位基因、血管紧张素转换酶（ACE）基因、激肽释放酶-激肽系统（KKS）通路的部分基因、可溶性鸟苷酸环化酶（sGC）基因等。     

大肠埃希菌连续分离株耐药表型及复方磺胺甲(噁)唑耐药相关基因研究

目的了解临床分离的大肠埃希菌耐药性和复方磺胺甲噁唑(SMZ/TMP)耐药相关基因存在状况。方法测定临床连续分离的60株大肠埃希菌对19种抗菌药物的敏感性,采用聚合酶链反应(PCR)检测SMZ/TMP耐药相关基因(sul1、dfrA1、dfrA17)。结果60株中sul1、dfrA1、dfrA17基因阳性分别为34株(56.7%)、1株(1.7%)、35株(58.3%)。结论临床分离的大肠埃希菌多重耐药严重;复方磺胺甲噁唑耐药相关基因携带率高。     

克拉霉素干混悬剂有关物质方法研究

目的 建立RP-HPLC法,测定克拉霉素干混悬剂的有关物质。方法 采用Agilent 1260 Infinity LC液相色谱仪,色谱柱为YMC C18(250mm×4.6mm, 5μm)柱,以0.033mol/L磷酸二氢钾溶液-乙腈(55:45)(用85%磷酸溶液或2mol/L氢氧化钾溶液调节pH值至5.4)为流动相,检测波长205nm,柱温50℃。结果 克拉霉素与9个已知杂质及其他未知杂质均能达到有效分离,克拉霉素及杂质E的线性范围分别为2~1000μg/mL(r=1.000)、0.9~6.75μg/mL(r=0.998);供试品溶液在室温25℃下的12h基本稳定。结论 本方法灵敏,准确,专属性强,重现性好,可用于克拉霉素干混悬剂有关物质的测定。     

桂西地区幽门螺杆菌对克拉霉素耐药的调查

目的:了解桂西地区幽门螺杆菌(HP)对克拉霉素的耐药情况.方法:对2006年6月～2007年12月行胃镜检查,并经尿素酶试验初步确认HP感染的患者.取其胃黏膜标本进行微需氧培养,经生化、涂片检查鉴定为典型的HP,经转代选择其中生长发育良好的菌株126株,应用K-B法药敏试验测定其对克拉霉素的敏感性.结果:126株HP菌株中,对克拉霉索耐药的有28株,耐药率为22.2%(28/126).结论:桂西地区HP菌株对克拉霉素耐药率略高,有必要定期检测或调查HP对抗生素的敏感性,以监测治疗的效果,提高根除HP的疗效.     

Effects of clarithromycin and verapamil on rabeprazole pharmacokinetics between CYP2C19 genotypes

Objective Rabeprazole as a proton pump inhibitor (PPI) is mainly reduced to rabeprazole thioether via a nonenzymatic pathway, with minor CYP2C19 and CYP3A4 involvement. The aim of this study was to compare possible effects of clarithromycin and verapamil as inhibitors of CYP3A4 on the pharmacokinetics of rabeprazole among CYP2C19 genotypes.Methods A three-way randomized, double-blind, placebo-controlled crossover study was performed. Nineteen volunteers, of whom six were homozygous extensive metabolizers (EMs), eight were heterozygous EMs, and five were poor metabolizers (PMs) for CYP2C19, received three 6-day courses of either daily 800 mg clarithromycin, 240 mg verapamil, or placebo in a randomized fashion, with a single oral dose of 20 mg rabeprazole on day 6 in all cases. Plasma concentrations of rabeprazole and rabeprazole thioether were monitored up to 24 h after the dosing.Results In the control phase, the AUC_0−∞ values for rabeprazole and rabeprazole thioether were 1,005±366 and 412±149 ng.h/ml in homozygous EMs, 1,108±340 and 491±245 ng.h/ml in heterozygous EMs, and 2,697±364 and 2,116±373 ng.h/ml in PMs, respectively. There were significant differences (p<0.001) in the AUC_0−∞ of rabeprazole and rabeprazole thioether among three different CYP2C19 genotypes. In the clarithromycin and verapamil phases, no significant differences were found in the pharmacokinetic parameters of rabeprazole compared with those in the control phase irrespective of CYP2C19 genotypes, whereas the AUC_0−∞ of rabeprazole thioether was significantly increased 2.8-fold and 2.3-fold in homozygous EMs (p<0.01), 2.0-fold and 2.0-fold in heterozygous EMs (p<0.05), and 1.6-fold and 1.9-fold in PMs (p<0.05), respectively. In each genotype group for CYP2C19, there were no statistical differences in the percent increase in those pharmacokinetic parameters between the clarithromycin and verapamil pretreatment phases.Conclusion The pharmacokinetic parameters of rabeprazole were not altered by clarithromycin or verapamil irrespective of the CYP2C19 genotypes. However, this result shows that both clarithromycin and verapamil significantly influence the disposition of rabeprazole by inhibiting the oxidation of the thioether, since the AUC_0−∞ of rabeprazole thioether that has no effect on acid secretion increased. Therefore, the pharmacokinetic interactions between rabeprazole and CYP3A4 or P-glycoprotein inhibitors have limited clinical significance.     

耐药肺结核患者结核分枝杆菌菌型分布及耐药研究

目的 研究肺结核耐药患者结核分枝杆菌菌型分布及耐药情况。方法 收集2011年4月至2012年7月肺结核患者痰液标本,分离培养后进行菌种鉴定和药敏试验。结果 人结核分枝杆菌约占83.91%,牛结核分枝杆菌约占15.70%;患者对一线抗结核药物耐药率显著高于对二线抗结核药物耐压率,差异有统计学意义(P<0.05);一类抗结核药物单一耐药率显著高于二类抗结核药物(χ²=4.281,P<0.05)。结论 肺结核耐药患者菌型以人结核分枝杆菌最多,对一线抗结核药物的耐药率显著高于对二线抗结核药物的耐药率。     

Metronidazole and clarithromycin resistance amongst Helicobacter pylori isolates from a large metropolitan hospital in the United States.

Background: Metronidazole and clarithromycin-based therapies are among the most efficacious treatment regimens for H. pylori infection. Resistance to metronidazole or clarithromycin is associated with impaired therapy with these agents. We conducted a retrospective review of susceptibility data to determine the frequency of primary metronidazole and clarithromycin resistance among H. pylori isolates from a single metropolitan hospital in the United States. The database comprised 933 patients who presented at the Digestive Diseases Clinic at the Veterans Affairs Medical Center in Houston between September 1988 and January 1997 with complaints of dyspepsia, abdominal pain and peptic ulcer disease. One hundred and seventy-nine of these patients had both pharmaceutical records available for evaluation and culture and antimicrobial susceptibility data for analysis. The MICs were determined by both E-test and broth microdilution tests. The frequency of primary metronidazole resistance was 37.4% (67/179). The level of primary clarithromycin resistance was 6.1%. Dual metronidazole and clarithromycin resistance was present in approx. 3%. The high level of primary metronidazole and clarithromycin resistance in H. pylori isolates from this metropolitan hospital is such that antimicrobial susceptibility data should be available so that informed choice can be made for specific eradication therapies, especially in patients who fail treatment.