Cutaneous manifestations of Nocardia brasiliensis infection in Taiwan during 2002–2012—clinical studies and molecular typing of pathogen by gyrB and 16S gene sequencing

To observe the clinicopathologic and resistance profiles of the Nocardia brasiliensis causing cutaneous nocardiosis in Taiwan, 12 N. brasiliensis isolates were prospectively collected from patients with cutaneous nocardiosis in a hospital during 2002–2012. Clinicopathologic data were obtained, and isolates were identified by biochemical methods and 16S rRNA sequencing. Susceptibilities to 14 antimicrobial compounds were tested. Isolates were further genotyped by sequencing of 16S rRNA, secA1, hsp65, and gyrB genes. The nodulopustular pyoderma associated with sporotrichoid spreading was the most common skin presentations caused by N. brasiliensis. All of the isolates were susceptible to amikacin, gentamicin, tobramycin, piperacillin/tazobactam, and trimethoprim/sulfamethoxazole and resistant to kanamycin, erythromycin, and oxacillin, while susceptibilities to imipenem, vancomycin, penicillin-G, tetracycline, clindamycin, and ciprofloxacin varied among the 12 isolates. GyrB genotyping delineated the 12 isolates into 2 major groups, which was coincident with different single nucleotide substitutions at position 160 (G versus T) of 16S rRNA, different levels of imipenem minimum inhibition concentration (4–32 versus 0.25–0.75 mg/L), and prevalence of lymphadenitis (66.7 versus 16.7%). We have noted that tiny pustular lesions can be the first sign of cutaneous nocardiosis, which we believe has not been previously emphasized. No resistance to trimethoprim and sulfamethoxazole was found; therefore, sulphonamide drugs remain effective for treatment of cutaneous nocardiosis in Taiwan.     

基于16S rRNA和gyrB基因的施万菌种水平鉴定分析

目的 构建基于16S rRNA和gyrB基因对施万菌(Shewanella)进行种水平鉴定的方法,比较2个基因的鉴定能力差异.方法 利用DnaSP 6.0软件对施万菌16S rRNA和gyrB基因的信息位点数及其百分比、核苷酸多态性值、平均G+C含量、非同义突变率与同义突变率的比值(Ka/Ks)、Tajima检验进行基...     

结核分枝杆菌对左氧氟沙星与莫西沙星的交叉耐药性及gyrA和gyrB基因突变分析

目的研究结核分枝杆菌(Mycobacterium tuberculosis,MTB)对左氧氟沙星(levofloxacin,LVF)与莫西沙星(moxifloxacin,MXF)的交叉耐药性,分析gyrA和gyrB基因突变位点分布及突变位点与耐药水平的关系。方法采用改良罗氏培养基检测MTB标准菌株H37Rv、66株LVF耐药和55株LVF敏感的MTB临床分离菌株LVF和MXF的最低抑菌浓度(minimal inhibition concentra-tion,MIC)。通过PCR直接测序法测定gyrA和gyrB耐药基因片段。结果 MXF的MIC比LVF低2~4倍,MXF抗MTB菌株的活性是LVF的2~4倍。MTB标准菌株H37Rv未见gyrA和gyrB基因突变。66株LVF耐药和55株LVF敏感菌株均存在gyrA AGC95ACC(Ser→Thr)突变;55株LVF敏感菌株gyrA和gyrB基因未见其他突变。66株LVF耐药菌株中,40株(60.6%)gyrAGAC94(AAC或GGC或GCC或CAC或TAC)(Asp→Asn或Gly或Ala或His或Tyr)突变;19株(28.8%)gyrA GCG90GTG(Ala→Val)和1株(1.5%)gyrA GCG90AAG(Ala→Lys)(未见报导)双碱基突变;3株(4.6%)gyrA TCG91CCG(Ser→Pro)突变;1株(1.5%)gyrB GAC500AAC(Asp→Asn)突变;2株(3.0%)呈gyrA GAC94(AAC或GCC)(Asp→Asn或Ala)与gyrB GGG551AGG(Gly→Arg)(未见报导)双位点突变。gyrA GAC94(AAC或GGC)(Asp→Asn或Gly)突变引起FQs药物较高水平耐药;GAC94GCC(Asp→Ala)和GCG90GTG(Ala→Val)突变引起FQs药物较低水平耐药。结论 LVF与MXF之间存在交叉耐药,MXF MIC随LVF MIC增高而增高,但耐药水平不同。GyrA基因突变位点可能与耐药水平有关,有可能根据基因突变的位点分析耐药水平。     

Geographically dependent distribution of gyrA gene mutations at codons 83 and 87 in Salmonella Hadar, and a novel codon 81 Gly to His mutation in Salmonella Enteritidis

In all, 90 nalidixic acid-resistant clinical strains of Salmonella Hadar and Salmonella Enteritidis isolated in Norway but of predominantly foreign origin were subjected to sequencing of the gyrA, gyrB, parC and parE genes. All the isolates contained at least one mutation in gyrA codon 83 or codon 87. A highly significant correlation between mutations in gyrA codon 83 and strains originating from Southeast Asia was found in S. Hadar but not in S. Enteritidis. A novel gyrA codon 81 Gly to His mutation was discovered in one S. Enteritidis isolate. One amino-acid (aa) changing mutation was found outside the quinolone resistance-determining region (QRDR) of S. Hadar parC at codon 57, which has previously only been observed once in Salmonellae.     

Salmonella enterica Serotype Typhi with nonclassical quinolone resistance phenotype

We report Salmonella enterica serotype Typhi strains with a nonclassical quinolone resistance phenotype (i.e., decreased susceptibility to ciprofloxacin but with susceptibility to nalidixic acid) associated with a nonsynonymous mutation at codon 464 of the gyrB gene. These strains, not detected by the nalidixic acid disk screening test, can result in fluoroquinolone treatment failure.     

短乳杆菌gyrB基因的扩增与测序

目的:为检测短乳杆菌gyrB基因序列。方法:设计两对简并引物对3种啤酒污染乳酸杆菌进行PCR扩增,获得短乳杆菌部分目的基因片断。并将该片断克隆至pGEM-T Easy载体,经限制性内切酶分析和测序获得短乳杆菌gyrB基因部分序列。结果:通过对序列进行进化树分析,发现该序列与啤酒典型污染菌植物乳酸杆菌gyrB序列相似性较高。结论:短乳杆菌gyrB序列对于建立啤酒中短乳杆菌快速检测具有十分重要的意义。     

A PCR-based method to differentiate between Acinetobacter baumannii and Acinetobacter genomic species 13TU

A new PCR-based method that exploits differences in gyrB gene sequences was developed to distinguish between Acinetobacter baumannii and Acinetobacter genomic sp. 13TU. Among 118 clinical and reference Acinetobacter strains, 102 of which were previously speciated by amplified rDNA restriction analysis as belonging to the Acinetobacter calcoaceticus-A. baumannii complex, the method correctly identified 31 A. baumannii and 54 Acinetobacter genomic sp. 13TU isolates to the species level. The method was rapid, specific and easy to interpret.     

GyrB基因在诺卡菌菌种鉴定中的应用研究

目的 分析研究gyrB基因对于诺卡菌种水平的鉴定能力。方法 采用特异性引物对14种41株诺卡菌的gyrB基因进行扩增,并对其产物进行测序。从GenBank数据库中下载21株诺卡菌gyrB基因序列,对序列用DNAStar软件计算其相似性。利用Mega 6.06软件,采用邻接法和最大似然法构建诺卡菌菌种系统发育进化树。结果 以gyrB基因构建的系统发育进化树中,不同诺卡菌种能准确地分离成独立的分支,且亲缘关系较近的近缘种也能得到有效分离。诺卡菌种gyrB基因种内相似性为94.2%~100%,种间相似性约为79.2%~97.6%,平均相似性为86.9%。结论 gyrB基因序列分析是一种快速、准确、特异性较高的鉴定方法,能将诺卡菌属的鉴定至种的水平,对于鉴定诺卡菌属近缘种gyrB基因比16S rDNA更具优越性。     

宁波地区耐多药结核分枝杆菌喹诺酮耐药gyr基因突变研究

目的 为阐明宁波地区耐多药结核分枝杆菌(Multiple drug-resistant tuberculosis, MDR-TB)的gyr基因突变特征,深入研究MDR-TB对喹诺酮类药物耐药与gyr基因突变特征的关系。方法 采用1%比例法对MDR-TB进行氧氟沙星药敏检测实验,通过 DNA直接测序法分析MDR-TB的gyr基因突变情况。结果 120株MDR-TB临床分离株中有34株对喹诺酮耐药,总耐药率为28.33%(34/120)。34株耐喹诺酮菌株中,30株gyr基因发生突变,突变率为88.24%(30/34)。30株gyr基因发生突变的菌株中gyrA基因突变有29株,占96.67%(29/30),突变位点包括90、91和94位氨基酸;gyrB基因突变有2株,其中1株均合并gyrA基因突变,占6.67%(2/30),突变位点包括499和502位氨基酸。结论 宁波地区MDR-TB对喹诺酮类药物耐药形势较为严峻,gyrA基因突变与MDR-TB对喹诺酮类药物耐药相关。     

副溶血弧菌的gyrB基因环介导的恒温扩增技术检测方法的研究

目的建立一种适合基层实验室应用和开展的快速检测副溶血弧菌的DNA环介导的恒温扩增法(LAMP)。方法针对副溶血弧菌的IgyrB/I 基因序列设计了4条引物（2条内引物、2条外引物）,并对扩增反应条件进行了优化。结果整个检测过程仅需1.5 h,可通过肉眼目测或电泳检 测判断结果;对3株种系背景明确的副溶血弧菌不同实验对照株、23株副溶血弧菌地方分离株和32株其他肠道菌进行了检测,具有很高的特异性 ;该方法的最低检测限为24 cfu/ml,具有良好的敏感性;应用于61份贝类海产品的现场检测,阳性率为100%,与实时荧光定量PCR方法的检测 结果相符,阳性率高于传统的培养鉴定方法。结论本方法具有快速、灵敏、特异、简便、经济等特点,适合基层实验室、应急检测或现场监测 等使用,具有较高的推广价值。