耐甲氧西林金黄色葡萄球菌检测方法应用评价

目的:评价四种检测耐甲氧西林金黄色葡萄球菌(MRSA)的实验方法.方法:收集107株金黄色葡萄球菌临床分离株,分别采用乳胶凝集实验法、头孢西丁纸片法、盐琼脂筛选法和聚合酶链反应(PCR)法进行实验检测.结果:PCR法是检测MRSA的金标准,特异性和灵敏度均为100%.头孢西丁纸片法和乳胶凝集法实验特异性均为100%,灵敏度是97.9%.盐琼脂筛选法特异性为91.4%,灵敏度为95.9%.结论:各级实验室需针对自身条件选择相应的实验方法,为临床提供准确的实验诊断.     

社区相关性耐甲氧西林金黄色葡萄球菌致病性研究进展

社区相关性耐甲氧西林金黄色葡萄球菌(CA-MRSA)自从上世纪八十年代出现以来,得到了全世界的普遍关注。CA-MRSA较强地适应外界环境的能力和定植能力对于其致病性起到了十分重要的作用,近期在CA-MRSA-USA300型菌株的基因组中发现的可移动精氨酸代谢元件(ACME),以及该段DNA编码的精氨酸脱亚胺酶(arginine deiminase)对MRSA在人体中的定植起到了重要作用。     

苏州地区耐甲氧西林金黄色葡萄球菌的临床分布特点及耐药性分析

目的 探讨苏州高新区人民医院耐甲氧西林金黄色葡萄球菌(MRSA)的临床分布特点及其耐药性,为临床治疗葡萄球菌属感染提供正确选药依据.方法 对2015年1月至2017年8月苏州高新区人民医院临床各科送检的各类标本中分离鉴定出的MRSA,采用MIC法进行药敏试验,并探讨其临床分布与耐药特点.结果 分离出的170株金黄色葡萄球菌中,MRSA有40株,占23.5%,以呼吸科病房为最多(20株),占50.0%、其次是重症监护病房(ICU)、儿科、神经内科;多重耐药的MRSA对所有β-内酰胺类高度耐药,对大环内酯类耐药率为67.5%;对克林霉素类耐药性较高,其中红霉素诱导的克林霉素耐药率为47.5%,提示该药在苏州地区已不能够用于MRSA 感染的治疗.对氨基糖苷类、氟喹诺酮类耐药率较低(耐药率<30%),磺胺甲噁唑、甲氧苄啶及利福平具有良好的活性,尚未发现耐糖肽类和利萘唑胺的菌株.结论苏州地区近3年来MRSA菌株检出率较低;对常用抗菌素呈多重耐药,尤其是对大环内酯-克林霉素抗菌药物耐药率高.因此应加强对MRSA耐药率的监测,合理选抗菌药物,从而减少MRSA 的产生和传播.     

脉冲场凝胶电泳分析耐甲氧西林金黄色葡萄球菌

目的探索一种准确、快速、可靠的分子生物学分型方法，研究耐甲氧西林的金黄色葡萄球菌（ＭＲＳＡ）的流行病学。方法建立脉冲场凝胶电泳（ＰＦＧＥ）的分子生物学分型方法。将细菌包埋于琼脂块中，原位溶解细菌，ＳｍａⅠ消化染色体ＤＮＡ，经脉冲场凝胶电泳（ＰＦＧＥ）分离，比较染色体限制性内切酶图谱，确定菌株的亲缘关系。结果３８株ＭＲＳＡ的ＰＦＧＥ谱分１０组（Ａ～Ｊ），以Ａ型为主，Ａ型又有Ａ１～Ａ４四个亚型。１９９５年１１月～１９９６年３月发生了Ａ１亚型株暴发流行，从１８位住院病人共分离到２２株。暴发流行株在住院病人、各病房及各医院之间进行传播。结论对ＭＲＳＡ的流行病学进行研究，ＰＦＧＥ是一种准确、可靠、重复性好的分子生物学分型方法。     

内在化的耐甲氧西林金黄色葡萄球菌诱导宿主细胞凋亡

目的 研究耐甲氧西林金黄色葡萄球菌(MRSA)的内在化过程及内在化后引起宿主细胞的变化情况，并探讨宿主细胞[Ca2 ]i变化与MRSA内在化及细胞凋亡之间的关系。方法 利用激光扫描共聚焦显微镜追踪宿主细胞[Ca^2 ]i的动态变化，流式细胞术观察细胞凋亡情况。结果 MBSA ATCC-67-O不能被Hep2细胞内在化，却可以进入ECV304．细胞，并导致ECV304细胞凋亡，抑制细胞[Ca^2 ]i的升高未能影响内在化的菌数和凋亡的细胞数。结论 内在化的MBSA ATCC-67-O能够诱导FLV304．细胞发生凋亡。宿主细胞[Ca2 ]i变化与MBSA ATCC-67-O内在化及细胞凋亡之间并非直接的因果关系。     

一起耐甲氧西林金黄色葡萄球菌医院感染的调查

目的分析一起耐甲氧西林金黄色葡萄球菌医院感染的危险因素,探索有效的控制措施。方法查看病房环境、回归性病例分析,利用PFGE方法对耐甲氧西林金黄色葡萄球菌（MRSA）菌株进行同源性分析。结果2007年6～7月MRSA感染9例,罹患率为15%;病房分布有聚集性;9份菌株药敏结果基本相同;5份菌株具有高度同源性;主要的危险因素有抗菌素的频繁使用、大面积的烧伤、与感染者同房、换床、陪护和探视等。结论合理使用抗生素,尽量减少侵袭性操作、严格隔离MRSA感染者、严格控制人员尤其探视人员进入、加强医护人员的手部卫生、加强器械及环境消毒、及时采样送检是预防和控制MRSA感染的关键。     

阴沟肠杆菌B8提取物对耐甲氧西林金黄色葡萄球菌的抑菌作用

阴沟肠杆菌Ｂ8分离自水稻叶片。临床分离菌株耐甲氧西林金黄色葡萄球菌 (ＭＲＳＡ) 87株 ,ＭＳＳＡ(不耐甲氧西林金黄色葡萄球菌 ) 38株 ,ＭＲＳＣＯＮ(耐甲氧西林凝固酶阴性葡萄球菌 ) 16株 ,ＭＳＳＣＯＮ(不耐甲氧西林凝固酶阴性葡萄球菌 ) 5株 ,大肠杆菌 10株 ,铜绿假单胞菌 2 5株 ,阴沟肠杆菌 14株 ,不动杆菌 2 6株 ,肺炎克雷伯菌 8株 ,共 2 2 9株。阴沟杆菌Ｏ2 9和Ｏ2 9Ｍ作标准菌株。将 3升ＬＢ固体培养基融化后制成平皿 ,将Ｂ8细菌培养液铺于平皿表面 ,30℃培养 48ｈ。刮去培养基表面菌体 ,将培养基切成约作者单位 :浙江大学医学院附…     

应用ED—PCR技术快速检测耐甲氧西林葡萄球菌

应用ＥＤ－ＰＣＲ技术从临床分离的７６株葡萄球菌中鉴定出耐甲氧西林金黄色葡萄球菌９株，耐氧西林血浆凝固酶阴性的葡萄球菌１１株。此结果与药物敏感性检测结果相符。ＥＤ－ＰＣＲ法具有简便，特异，省时，耗资少等优点，宜在临床检验室推广应用。     

耐甲氧西林金黄色葡萄球菌的分子生物学研究进展

耐甲氧西林金黄色葡萄球菌(Methicillin-resistant Staphylococcus aureus，MRSA)不仅对所有β-内酰胺类抗生素具有内在耐药性，而且通过获得其它耐药基因而呈现对多数其它类抗菌药的多重耐药性，给临床治疗带来严重问题。本从分子生物学水平就MRSA耐药决定因子(mec)与辅助基因(fem)在耐药性产生中的作用作一简要综述。     

检测耐甲氧西林葡萄球菌及其耐药性分析

为了解我院耐甲氧西林葡萄球菌（MRS）的感染情况及其耐药性,为临床治疗MRS感染提供实验室依据和有效的用药措施．用微生物鉴定仪检测了142株葡萄球菌对13种抗生素的MIC,对苯唑青霉素MIC＞2ug／ml的菌株,采用K－B纸片法,补充检测对苯唑青毒素纸片的耐药性,共筛选出80株MPS,其中耐甲氧西林金黄色葡萄球菌（MRSA）20株,耐甲氧西林表皮葡萄球菌（MRSE）33株,其它耐甲氧西林葡萄球菌27株,说明感染我院的MRS种类不断增加,应引起重视．药散结果表明MRS对万古霉素（Van）100％敏感,对其他抗生素的敏感性明显低于对甲氧西林敏感的葡萄球菌（MSS）,因此,临床上应区分MRS与MSS,选择性地用药．     

High prevalence of class 1 integrons in clinical isolates of methicillin-resistant Staphylococcus aureus from India

Introduction: Class1 integrons are one of the prevalent mechanisms of antibiotic resistance gene transfer in Gram-negative organisms, but their prevalence and role in the spread of antibiotic resistance genes in methicillin-resistant Staphylococcus aureus (MRSA) is unexplored. The purpose of this study was to investigate the prevalence of class 1 integrons in clinical isolates of MRSA. Materials and Methods: Total 143 MRSA isolates obtained from two different cities in India (Pune and Mumbai) were characterized by biochemical tests, and the antibiotic sensitivity was performed using the Clinical and Laboratory Standards Institute (CLSI) guidelines. The presence of class 1 integrons, sul1/qacEΔ1 region of class 1 integron and mecA gene among these isolates was determined by polymerase chain reaction (PCR). Results: All 143 isolates were mecA positive and coagulase-positive. Overall, 71% of the MRSA isolates carried class 1 integrons; 58% (45/77) of the isolates obtained from Mumbai and 85% (56/66) of the isolates from Pune carried class 1 integrons. In all, 39% of these isolates carried sul1/qacEΔ1 region, thus confirming the association of class 1 integrons with antibiotic resistance genes. Along with β-lactam antibiotics the MRSA isolates were resistant to several other antibiotics, with resistance to erythromycin, ciprofloxacin and trimethoprim-sulfamethoxazole being observed in 75%, 66% and 60% of the isolates, respectively. Conclusion: To the best of our knowledge, this is the first report of class 1 integrons in MRSA isolates from India. The study provides insights into the prevalence of a novel mechanism adapted by MRSA for the propagation of antibiotic resistance genes.     

Increasing incidence and widespread dissemination of methicillin-resistant Staphylococcus aureus (MRSA) in hospitals in central Europe, with special reference to German hospitals

Objective: to present data on prevalence and interregional spread of methicillin-resistant Staphylococcus aureus (MRSA) in Germany.
Methods: A nationwide collection of MRSA isolates from nosocomial infections in 143 hospitals was established from isolates ( n =4368) sent to a microbiological reference center during 1993–95. As chosen by distinguishable resistance phenotypes at each time of occurrence during the study period, 1830 isolates were subjected to molecular typing by means of Sma l macrorestriction patterns, PCR for RNA gene spacer patterns, and PCR for patterns of DNA stretches flanked by the ERIC-2 sequence and flanked by Tn 976 and ribosomal binding site. In addition, data from a multicenter study on the incidence of antibiotic resistance have been analyzed (32 centers, 637 S. aureus isolates).
Results: In 1995 the prevalence of MRSA among S. aureus isolates was 8.7% overall in central Europe (including Germany), in comparison to 1.7% in 1990. From 1993 until now, a continuous interregional dissemination of six epidemic strains, which were identified by molecular typing, was recorded. Besides these epidemic strains, 15 MRSA strains were identified which could not be allocated to the epidemic MRSA or to the known clonal groups of the species S. aureus. MRSA from three cases of sporadic nosocomial infections exhibited characteristics of the clonal group of S. aureus with the capacity for toxic shock syndrome formation. The pattern of one MRSA corresponded to those of the S. aureus group exhibiting phage pattern 94,96.
Conclusions: The prevalence of MRSA has increased in central Europe (and Germany) during the last 5 years, to 8.7%. The main source of infection with MRSA is obviously interregional dissemination of epidemic strains. At the same time, the mecA gene has been acquired by strains previously sensitive to methicillin.     

Predictors of clinical and microbiological treatment failure in patients with methicillin-resistant Staphylococcus aureus (MRSA) bacteraemia: a retrospective cohort study in a region with low MRSA prevalence

Invasive infections with methicillin-resistant Staphylococcus aureus (MRSA) have been associated with increased morbidity and mortality. The aim of the present study was to identify independent predictors of early mortality and treatment failure in patients with MRSA bacteraemia. A total of 132 adult patients who developed MRSA bacteraemia during hospitalization in the University Hospital of Vienna between 2000 and 2011 were screened and 124 were included in a retrospective cohort study. Patient demographics, source of bacteraemia, antimicrobial treatment and microbiological characteristics were evaluated. The 28-day crude mortality was 30.6%. Predictors of early mortality identified in multivariate Cox regression analysis included higher patientage (adjusted hazard ratio (aHR) 1.03, 95% CI 1.01–1.06, p 0.006), pneumonia (aHR 3.86, 95% CI 1.83–8.12, p <0.001) and failure to use MRSA active treatment (aHR 8.77, 95% CI 3.50–21.93, p <0.001). Ninety-one (73.4%) patients received glycopeptides as specific MRSA treatment. Of 63 patients treated with vancomycin, only 14 (22.6%) patients had aimed trough levels of 15–20 mg/L. Vancomycin MIC ≥ 2 mg/L was detected in 28.2% and was associated with glycopeptide pretreatment (p 0.001). All MRSA isolates were susceptible to linezolid and tigecycline. Persistent bacteraemia ≥ 7 days was documented in 25 (20.2%) patients. Independent determinants for microbiological eradication failure in patients with MRSA bacteraemia included endocarditis (p <0.001) and vancomycin trough levels (p 0.014), but not vancomycin MIC. Failure of clinical and microbiological eradication of MRSA among patients with MRSA bacteraemia was associated with clinical entity rather than with bacterial traits. Pharmacokinetic parameters seem to be decisive on microbiological and clinical success.     

肝移植术后耐甲氧西林金黄色葡萄球菌血流感染危险因素的17年回顾性分析

目的探讨本院肝移植术后耐甲氧西林金黄色葡萄球菌（MRSA）血流感染的危险因素及临床结果。方法回顾分析1993年1月至2010年5月肝移植术后血液MRSA阳性感染者的资料。结果 695例肝移植患者中,53例（7.6%）出现革兰氏阳性球菌血流感染,以肠球菌最为常见,7例病人发生7次MRSA血流感染。分析肝移植术后MRSA血流感染的危险因素发现,急性排斥（P=0.03）是出现MRSA血流感染的危险因素。肝移植术后MRSA血流感染与非MRSA血流感染的15d、30d、1年病死率差异无统计学意义。结论急性排斥是出现肝移植术后MRSA血流感染的危险因素,但肝移植术后MRSA血流感染后15d、30d、1年的死亡率未明显增加。     

High diversity of Panton–Valentine leukocidin-positive, methicillin-susceptible isolates of Staphylococcus aureus and implications for the evolution of community-associated methicillin-resistant S. aureus

In total, 100 Staphylococcus aureus isolates from diverse cases of skin and soft-tissue infection at a university hospital in Saxony, Germany, were characterised using diagnostic microarrays. Virulence factors, including Panton-Valentine leukocidin (PVL), were detected and the isolates were assigned to clonal groups. Thirty isolates were positive for the genes encoding PVL. Only three PVL-positive methicillin-resistant S. aureus (MRSA) isolates were found, two of which belonged to European clone ST80-MRSA IV and one to USA300 strain ST8-MRSA IV. The remaining methicillin-susceptible PVL-positive isolates belonged to a variety of different multilocus sequence types. The predominant strains were agrI/ST22, agrII/CC5, agrIII/CC30 and agrIV/ST121. In order to check for possible bias caused by regional or local outbreak strains, an additional 18 methicillin-susceptible, PVL-positive isolates from the UK were tested. Approximately two-thirds of the UK isolates belonged to types that also comprised approximately two-thirds of the isolates from Saxony. Some methicillin-susceptible PVL-positive isolates (agrI/ST152, agrIII/ST80 and agrIII/ST96) closely resembled known epidemic community-acquired MRSA (CaMRSA) strains. These findings indicate that the current rise in PVL-positive CaMRSA could be caused by the dissemination of novel SCCmec elements among pre-existing PVL-positive strains, rather than by the spread of PVL phages among MRSA strains.     

金黄色葡萄球菌肠毒素A基因的克隆及原核表达

目的从分离培养的金黄色葡萄球菌中提取肠毒素A（SEA）基因,亚克隆入表达载体pET-28a（＋）,诱导融合蛋白的表达,为肠毒素A应用研究奠定基础。方法根据GenBank中SEA的序列,设计一对特异性引物,以金黄色葡萄球菌DNA为模板PCR扩增SEA,纯化DNA进行BamHⅠ、XholⅠ双酶切鉴定及测序鉴定,并与做相应酶切的pET-28a（＋）连接,转化大肠杆菌BL21,并对质粒进行双酶切鉴定及基因序列分析,用IPTG诱导融合蛋白的表达,His标签单克隆抗体进行免疫印迹验证融合蛋白的表达。结果以金黄色葡萄球菌DNA为模板,成功扩增了SEA基因,基因大小为774bp,重组PET-28a（＋）-SEA双酶切鉴定可见目的片段,测序结果显示SEA在正确读框中,序列比对分析显示其与相关报道核苷酸序列一致性达99.9%。经IPTG诱导后,SDS-PAGE可见pET-28a（＋）-SEA/BL21在相应分子量（约30000Mr）条带大量表达,免疫印迹能检测到目的蛋白条带,说明其与His标签融合表达。结论克隆了金黄色葡萄球菌SEA基因,并成功在大肠杆菌BL21中融合表达,为肠毒素A应用研究奠定了基础。     

Evaluation of three chromogenic media (MRSA-ID, MRSA-Select and CHROMagar MRSA) and ORSAB for surveillance cultures of methicillin-resistant Staphylococcus aureus

Screening specimens were homogenised in saline 0.9% w/v before either direct inoculation or following enrichment in broth on three chromogenic media (MRSA-ID, CHROMagar MRSA and MRSA Select) and ORSAB medium for the detection of methicillin-resistant Staphylococcus aureus (MRSA). In total, 102 of 466 specimens yielded MRSA on at least one medium. After incubation for 16-18 h, the sensitivity was 51%, 59%, 47% and 65% on MRSA-ID, CHROMagar MRSA, ORSAB and MRSA Select, respectively, compared with 82%, 75%, 67% and 80%, respectively, after 42 h, and 93%, 95%, 79% and not tested, respectively, following broth enrichment. There were significantly more MRSA colonies on MRSA-Select after 16-18 h than on ORSAB or MRSA ID (p 0.001 and 0.0022, respectively), whereas there were more MRSA colonies after 42 h on MRSA-ID and MRSA-Select than on ORSAB (p 0.0004 and 0.012, respectively). The specificity of the media for identifying MRSA based on the colour of colonies after incubation for 16-18 h was 100%, 99%, 99% and 100%, respectively, compared with 98%, 97%, 98% and 98%, respectively, after 42 h, and 100%, 99%, 100% and not tested, respectively, following broth enrichment. The speed of detection (mean time to report a positive result) was 1.65, 1.72, 2.31 and 1.35 days, respectively. For each of the three media tested following enrichment, the use of an enrichment broth increased the detection rate of MRSA by 16-24%.