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目的 比较脉冲场凝胶电泳(PFGE)和多位点串联重复序列分析(MLVA)分型方法用于我国肠炎沙门菌分子分型的能力,建立我国肠炎沙门菌MLVA分型标准操作方法及数据库。方法 根据国际PulseNet公布的肠炎沙门菌PFGE和MLVA分型方案,对来自我国6个省(直辖市)的289株肠炎沙门菌进行分子分型分析,并结合流行病学资料,评价这两种分型方法对我国肠炎沙门菌分离株的分型能力。结果 289株肠炎沙门菌经XbaⅠ酶切,PFGE后获得55种带型,其分辨能力(D值)为0.8433。PFGE优势带型为JEGX01.CN0003及JEGX01.CN0001,带型频率分别为35.6%、25.6%,但二者仅表现两个条带的差异,其余型别均低于5%。采用MLVA分析,获得63种型别,分为2个群,D值为0.8608,说明MLVA分辨能力高于单酶切PFGE,但分型能力仍然有限。MLVA主要型别ST1包含了97株菌株,占37.5%,分布于各省及各年份。若联合PFGE及MLVA分型,则产生104种型别,D值为0.9058。对流行病学调查显示为肠炎沙门菌暴发病例菌株进行PFGE双酶切及MLVA分型,结果均显示这些菌株具有明显的聚集性,但不能与同期散发菌株完全区分开。结论 MLVA与PFGE分型方法的分辨能力在肠炎沙门菌中较低,在确认肠炎沙门菌引起的暴发事件时,需紧密结合流行病学调查资料,采用双酶切PFGE或MLVA进行分型分析。 相似文献
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目的 研究海南省类鼻疽伯克霍尔德菌核糖体16S-23S内转录间隔区(ITS)序列多态性及基因型特征,了解该省与其他类鼻疽流行区菌株间的遗传背景差异。方法 PCR扩增272株类鼻疽伯克菌的ITS片段并通过毛细管凝胶电泳检测产物长度;通过DNA测序及序列比对确认不同长度ITS的基因型及序列一致性;通过2检验分析不同流行区ITS型别的分布差异。结果 经毛细管凝胶电泳及DNA测序确认,272株类鼻疽伯克菌中发现C、E、CE、G 4种ITS基因型:C型64株(23.53%),E型144株(52.94%),CE型56株(20.59%),G型8株(2.94%)。序列比对证实,海南省类鼻疽伯克菌中各型别(C、E、G)ITS序列高度保守,不同型别ITS的长度变化由其主要变异区的序列差异引起。我国海南省与泰国流行区的ITS型别分布差异有统计学意义(2=8.296,P0.05),与澳大利亚流行区分布差异无统计学意义(2=5.521,P0.05)。结论 海南省类鼻疽伯克菌临床菌株中存在C、E、CE、G 4种ITS基因型,C、E、CE为优势型别;与泰国、澳大利亚等传统流行区相比,其G型菌株比例较高。 相似文献
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目的应用分子分型技术对2007年上海和重庆市沙门菌监测点的50株汤卜逊沙门菌进行分子流行病学分析和抗生素敏感性测定,了解上海和重庆两地菌株的分子分型特征和药物敏感性特征。方法抗生素敏感性测定采用微量肉汤稀释法,分子分型方法包括脉冲场凝胶电泳(PFGE)和多位点串联重复序列分析(MLVA)。结果药敏试验显示78%的菌株存在多重耐药,其中磺胺甲恶唑和四环素的耐药率最高,其次是甲氧苄胺嘧啶;对头孢类抗生素(头孢噻肟、头孢三嗪、头孢他啶、头孢噻呋)均未产生耐药性。PFGE将50株菌分为15个带型,30株重庆分离株间有较高的相似性;MLVA分析显示除Sal16位点外,其余检测位点在所有待检菌株中没有差别。结论分子分型支持汤卜逊沙门菌引起的暴发以及散发。目前MLVA分型应用于汤卜逊沙门菌分子分型时,分型能力低于PFGE,需要进一步优化。 相似文献
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目的 比较脉冲场凝胶电泳(PFGE)和两种国际多位点串联重复序列分析(MLVA)分型方法用于我国鼠伤寒沙门菌分子分型的能力,并初步确定适用于我国鼠伤寒沙门菌MLVA分型中的VNTR位点。
方法 根据国际PulseNet公布的沙门菌PFGE分型方案、MLVA分型方案(包含7个VNTR位点,简称MLVA_PN)及欧洲食源性疾病监测网公布的鼠伤寒沙门菌MLVA分型方案(包含5个VNTR位点, 简称MLVA_EU),对来自我国5个省(直辖市)的175株鼠伤寒沙门菌进行分子分型分析,并结合流行病学资料,评价这三种分型方法对我国分离的鼠伤寒沙门菌的分型能力。
结果 175株鼠伤寒沙门菌经XbaⅠ酶切,PFGE后,获得56种带型,其分辨能力(D值)为0.8823。对其中的主优势带型JPXX01.CN0001菌株55株进一步用第二种内切酶BlnⅠ酶切后,获得6种带型。用MLVA_EU分型,获得96种型别,D值为0.9758。应用MLVA_PN分析,获得98种型别, D值为0.9763。 两种MLVA分型方法对菌株的分辨能力几乎相同,且分型结果具有较高的一致性。对流行病学调查显示为鼠伤寒沙门菌暴发患者和食物来源的菌株进行PFGE双酶切及MLVA分型,三种分型方法获得一致性结果,均显示这些菌株具有明显的聚集性。
结论 两种MLVA分型方法的分辨能力均高于PFGE,在确认鼠伤寒沙门菌引起的暴发事件时,采用需时较短,操作更方便的5个VNTR位点的MLVA分型方法可满足菌株聚集性分析。 相似文献
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目的评估不同培养基生长类鼻疽伯克霍尔德菌在Vitek 2 Compact中的鉴定结果。方法收集2010年6月至2017年5月海南省人民医院分离的类鼻疽伯克霍尔德菌127株,分别接种于哥伦比亚羊血平板(CBA)、麦康凯平板(MAC)和流感嗜血巧克力平板(CHA)上,以多位点序列分型(MLST)为金标准,评估Vitek 2 Compact VT2.R7.01版GN卡对不同培养基上生长的类鼻疽伯克霍尔德菌的鉴定准确性。结果 CBA平板上生长的类鼻疽伯克霍尔德菌鉴定准确率最高,为98.4%,2株被错误鉴定为洋葱伯克霍尔德菌;MAC次之,准确率为94.5%,其中4株被错误鉴定为洋葱伯克霍尔德菌,2株为伯克霍尔德菌某种,1株被错误鉴定为铜绿假单胞菌;CHA鉴定准确率最低,为91.3%,其中4株被错误鉴定为洋葱伯克霍尔德菌,2株为伯克霍尔德菌某种,5株无鉴定结果。结论 Vitek 2 Compact GN卡对不同培养基上类鼻疽伯克霍尔德菌鉴定准确率不同,建议优先选择CBA平板上菌落。 相似文献
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目的 评估不同培养基上类鼻疽伯克霍尔德菌(Burkholderia pseudomallei,BP)Vitek 2 compact的鉴定结果。方法 收集2010年1月-2017年5月间海南省人民医院类鼻疽伯克霍尔德菌127株,分别将其接种于三种不同培养基上,以多位点序列分型(MLST)为金标准,评估不同培养基上生长的类鼻疽伯克霍尔德菌,采用Vitek 2 compact VT2.R7.01版的 GN卡进行鉴定并比较其准确性。结果 哥伦比亚羊血平板(CBA)上的类鼻疽伯克霍尔德菌鉴定准确率最高,为98.4%,2株被鉴定为洋葱伯克霍尔德菌;麦康凯平板(MAC)次之,准确率为94.5%,其中4株被错误鉴定为洋葱伯克霍尔德菌,2株无法鉴定到种,1株被错误鉴定为铜绿假单胞菌;流感嗜血巧克力平板(CHA)鉴定准确率最低,为91.3%,其中4株被错误鉴定为洋葱伯克霍尔德菌,3株无法鉴定到种,4株鉴定不出结果。结论 该研 相似文献
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Szczypa K Sadowy E Izdebski R Hryniewicz W 《The Journal of antimicrobial chemotherapy》2004,54(4):828-831
OBJECTIVES: The aim of this study was to investigate Polish clinical isolates of Streptococcus pyogenes collected during a 7 year period using phenotypic and genotypic techniques. METHODS: A total of 816 isolates of S. pyogenes recovered from 33 medical centres in Poland were tested for their susceptibility to various antimicrobial agents. Erythromycin-resistant isolates were analysed by PFGE, multilocus sequence typing and emm typing methods. RESULTS: The tetracycline resistance rate was high (43%) among all S. pyogenes strains. Ninety-eight (12%) isolates were resistant to erythromycin. A low prevalence of the M phenotype (5.1%) associated with the presence of the mef(A) gene was found. All the isolates of the iMLSB phenotype harboured the erm(TR) gene. Out of the cMLSB isolates, 71.4% and 28.6% carried erm(TR) and erm(B), respectively. All isolates with erm(B) were resistant to telithromycin. PFGE analysis discerned 13 different patterns, A-N, with two predominant PFGE profiles--A (41 isolates) and B (25 isolates)--that in multilocus sequence typing corresponded, respectively, to a novel sequence type (ST) 367 and ST63. Overall, the representatives of these clones accounted for >90% of isolates of the iMLSB phenotype. CONCLUSIONS: A significant increase in erythromycin resistance was observed among clinical S. pyogenes collected in Poland over a 7 year period driven by the spread of two epidemic clones. 相似文献
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目的分析围产期李斯特菌感染6例的临床及其分子流行特征,为围产期李斯特菌感染的治疗、预防和控制提供参考依据。方法回顾性分析李斯特菌母婴感染病例的临床信息,对分离的病原菌进行血清学分型、脉冲场凝胶电泳分析、多位点序列分型及毒力基因检测。结果 6例孕妇出现发热、腹痛、胎动异常等症状;2例胎死宫内,4例新生儿出现胃肠炎、血流感染和脑膜炎等感染症状,其中1例新生儿出生2 d后死亡。6株产单核细胞李斯特菌有3种血清型(4b、1/2b和1/2a)、4种ST型(ST1、ST2、ST5和ST91)和5种PT型(pulsotype)。毒力基因(plcB、actA、hly、iap、prfA、inlA)检测均为阳性。结论 6例围产期李斯特菌感染病例症状较为严重,胎儿/新生儿预后差。分离的产单核细胞李斯特菌以血清型4b型和ST2型为主,携带6个毒力基因,致病力强。 相似文献
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Lee SH Chong CE Lim BS Chai SJ Sam KK Mohamed R Nathan S 《Diagnostic microbiology and infectious disease》2007,58(3):263-270
Burkholderia pseudomallei is a Gram-negative saprophytic soil bacterium, which is the etiologic agent of melioidosis, a severe and fatal infectious disease occurring in human and animals. Distinct clinical and animal isolates have been shown to exhibit differences in phenotypic trait such as growth rate, colony morphology, antimicrobial resistance, and virulence. This study was carried out to gain insight into the intrinsic differences between 4 clinical and 6 animal B. pseudomallei isolates from Malaysia. The 16S rRNA-encoding genes from these 10 isolates of B. pseudomallei were sequenced to confirm the identity of these isolates along with the avirulent Burkholderia thailandensis. The nucleotide sequences indicated that the 16S rRNA-encoding genes among the 10 B. pseudomallei isolates were identical to each other. However, the nucleotide sequence differences in the 16S rRNA-encoding genes appeared to be B. pseudomallei and B. thailandensis specific. The growth rate of all B. pseudomallei isolates was determined by generating growth curves at 37 degrees C for 72 h. The isolates were found to differ in growth rates with doubling time varying from 1.5 to 2.3 h. In addition, the B. pseudomallei isolates exhibited considerable variation in colony morphology when grown on Ashdown media, brain-heart infusion agar, and Luria-Bertani agar over 9 days of observation. Antimicrobial susceptibility tests indicated that 80% of the isolates examined were Amp(R) Cb(R) Kn(R) Gm(R) Chl(S) Te(S). Virulence of the B. pseudomallei clinical and animal isolates was evaluated in B. pseudomallei-susceptible BALB/c mice. Most of the clinical isolates were highly virulent. However, virulence did not correlate with isolate origin since 2 of the animal isolates were also highly virulent. 相似文献
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DiversiLab系统鲍曼不动杆菌基因分型可行性研究 总被引:2,自引:0,他引:2
目的 应用DiversiLab系统对鲍曼不动杆菌进行基因分型,并评价其可行性.方法 对2005年全国15个城市医院临床分离的58株鲍曼不动杆菌,应用以rep-PCR为原理的DiversiLab系统分型技术进行分型,并与脉冲场凝胶电泳分型和多位点序列分型技术结果进行比较;计算Simpson相关系数(Simpson's index of diversity)比较3种方法的分辨力.结果 DiversiLab系统将58株鲍曼不动杆菌分为5簇流行克隆和25个散发克隆;多位点序列分型技术将其分为35个序列型,并归为一个克隆复合体CC22(Clone Complex 22)和35个单一体(singleton);PFGE将其分成5个流行脉冲型和34个散发克隆.DiversiLab系统、MLST和PFGE的Simpson相关系数分别为0.876、0.944和0.961.结论 DiversiLab系统的操作简单、快速,可重复性好,可以作为大量菌株首选分型方法,但分辨力低于PFGE和MLST两种分型技术. 相似文献
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Lee YT Lin DB Wang WY Tsao SM Yu SF Wei MJ Yang SF Lu MC Chiou HL Chen SC Lee MC 《Diagnostic microbiology and infectious disease》2011,70(2):175-182
We used molecular typing methods to investigate an outbreak of methicillin-resistant Staphylococcus aureus (MRSA) infections in a respiratory care ward in Taiwan. From March to June 2006, the incidence of MRSA infection increased 3.75-fold. The overall carrier rates among the health care workers (HCWs) were 31.3% (total S. aureus), 16.4% (MRSA), and 14.9% (methicillin-sensitive SA, MSSA). Pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST), antibiograms derived from susceptibility testing of MRSA isolates, and multiplex polymerase chain reaction (PCR) provided strong epidemiologic and microbiologic evidence that the outbreak of MRSA infections at our hospital was linked to the same PFGE pulsotype A SCCmec type II, pvl-negative, MLST ST5 strain of MRSA isolated from seven HCWs and five patients. The outbreak was controlled by application of topical fucidin ointment to the anterior nares in all colonized HCWs. Multiplex PCR combined with PFGE and MLST is a feasible method for outbreak investigations in routine clinical laboratories. 相似文献
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Heym B Le Moal M Armand-Lefevre L Nicolas-Chanoine MH 《The Journal of antimicrobial chemotherapy》2002,50(3):323-329
Infection by methicillin-resistant Staphylococcus aureus (MRSA) presents a serious problem, as these organisms are resistant to a wide range of antibiotics. Moreover, MRSA with reduced susceptibility to glycopeptides (GISA) have emerged in recent years. In our hospital we were faced with an outbreak of GISA between January and March 2000, involving six patients who were free of MRSA when the GISA isolate emerged, and five of whom had not been given glycopeptides. The initial GISA isolate from the six patients was compared with six 'historical' multiple-resistant MRSA isolates (1996-1999), which had not been found to be GISA by the routinely used method. Antibiotic susceptibility was studied through the disc diffusion method, and MICs of glycopeptides were determined by Etest, agar and broth dilution techniques. Molecular strain typing was done by PFGE and multilocus sequence typing (MLST). All 12 isolates that belonged to the gentamicin-resistant group of MRSA were susceptible to vancomycin, but showed reduced susceptibility to teicoplanin through at least one MIC method. PFGE typing yielded five different but closely related profiles, and eight of the 12 clinical isolates were classified into a single PFGE type. MLST yielded an identical allelic profile for all 12 isolates, corresponding to the profile (3-3-1-12-4-4-16) of the 'Iberian' clone HPV107 of MRSA. MLST allowed us to confirm the presence of the 'Iberian' MRSA clone HPV107 with the same allelic profile in our hospital for several years. We can now note that strains of this clone have acquired reduced susceptibility to teicoplanin. 相似文献
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Marian Garcia-Nuñez Sara Quero Stella Catini Maria Lluisa Pedro-Botet Lourdes Mateu Nieves Sopena Miguel Sabria 《Journal of infection and chemotherapy》2013,19(5):896-901
An outbreak of Legionnaires’ disease with 113 confirmed cases was reported in the town of Mataró, Spain, in August 2002. In this study, we compared three different typing methods and characterized the clinical isolates by comparing them with other clinical isolates with the same ST from our own database to further characterize the outbreak. In the outbreak, a total of 16 clinical (nine patients) and 32 environmental (from four environmental sources) Legionella pneumophila isolates were analyzed by pulsed-field electrophoresis (PFGE), sequence-based typing (SBT), and monoclonal antibody typing (MAb). We compared the MAb and SBT profiles of the outbreak clinical isolates and other unrelated clinical isolates showing the same ST profile. We obtained seven different PFGE and SBT profiles and six MAb patterns from the outbreak isolates. PFGE and SBT showed 100 % concordance during the outbreak. SBT proved to be highly discriminatory, particularly with the addition of the new neuA gene. One PFGE, SBT (ST-37), and Philadelphia profile was observed among the clinical isolates. Using PFGE, this ST37 Philadelphia profile was closely related to other unrelated clinical isolates. These findings suggest that the ST37 Philadelphia profile could be a virulence marker in our area. The combination of the three methodologies was useful to further characterize and obtain additional information on a very explosive outbreak. Despite the minor discrimination of PFGE versus SBT, the two genetic methods are recommended in outbreak investigations. Further studies are currently underway in this area to obtain more definitive conclusions. 相似文献
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Pseudomonas pseudomallei, a common pathogen in Thailand that is resistant to the bactericidal effects of many antibiotics. 总被引:1,自引:1,他引:1 下载免费PDF全文
T Sookpranee M Sookpranee M A Mellencamp L C Preheim 《Antimicrobial agents and chemotherapy》1991,35(3):484-489
The purpose of this investigation was to identify newer antimicrobial agents that may be useful in the therapy of melioidosis. The in vitro susceptibilities of 199 clinical isolates of Pseudomonas pseudomallei to 22 antibiotics were determined by standard disk diffusion, and those to 13 antibiotics were determined by agar dilution. Over 90% of the isolates were susceptible to imipenem, piperacillin-tazobactam, piperacillin, ceftazidime, ticarcillin-clavulanate, ampicillin-sulbactam, and carumonam by both methods. Standard disk diffusion yielded unacceptably high false-susceptibility results with aztreonam, ciprofloxacin, and temafloxacin. Piperacillin, ceftazidime, imipenem, and ciprofloxacin were not bactericidal for three selected P. pseudomallei strains as determined by time-kill curve methods. Furthermore, addition of ciprofloxacin to piperacillin, ceftazidime, or imipenem did not enhance bactericidal activity. One hundred ninety-four strains showed weak beta-lactamase production that did not increase upon incubation with cefoxitin. These findings suggest that several newer antimicrobial agents may prove useful in the treatment of melioidosis. However, results of susceptibility studies involving P. pseudomallei and newer agents must be interpreted with caution. 相似文献
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Francis A Aiyar S Yean CY Naing L Ravichandran M 《Diagnostic microbiology and infectious disease》2006,55(2):95-99
Isolation and culture of Burkholderia pseudomallei remains the main stay in the diagnosis of melioidosis. Thus, the search for selective and differential media for B. pseudomallei has been ongoing. A number of such media have been reported with varying efficacy. Ashdown medium is the most established selective medium for the isolation of B. pseudomallei. There are no reports of differential media differentiating B. pseudomallei from Burkholderia cepacia. This report documents such a selective and differentiating medium for B. pseudomallei. Of a total of 1042 clinical specimens containing mixed flora and gram-negative isolates that were tested on this medium, 16 of the specimens yielded B. pseudomallei. The isolation rate was found to be 1.5%. This medium was found to be simple and inexpensive, can be made by small laboratories, and called as Francis medium. Based on the colony morphology and color, a preliminary report can be made within 18-24 h for the presence of B. pseudomallei. Our study showed that this medium had an overall sensitivity of 78.4% with a specificity of 92.2%. The use of this medium as an early diagnostic tool will help to reduce mortality and morbidity of melioidosis patients. 相似文献
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Oughton M Dascal A Laporta D Charest H Afilalo M Miller M 《Diagnostic microbiology and infectious disease》2011,69(2):213-217
To analyze 67 clinical methicillin-resistant Staphylococcus aureus (MRSA) isolated from pediatric hospital infections, we used multilocus variable-number tandem-repeat DNA sequence-based techniques, targeting the protein A polymorphic X region and the clumping factor B complete R domain. We define a "clfB similarity score" and then compare the double loci analysis of closely related MRSA isolates with pulsed-field gel electrophoresis (PFGE). We found an endemic clone (MLST-ST8, spa-t008, SCCmecIV, ClfB lineage 1) able to originate 3 possible outbreaks and a second clone (MLST-ST152, spa-t355, SCCmecV, ClfB lineage 4) responsible for limited cases of MRSA infections, indicating that the combination of spa and clfB-lineage typing is useful to trace MRSA pediatric outbreaks. 相似文献