鲍氏不动杆菌耐喹诺酮类药物的机理研究

目的：研究鲍氏不动杆菌对喹诺酮类药物的耐药机制。方法：对临床上收集的耐环丙沙星鲍氏不动杆菌进行氟罗沙星摄取试验、外膜蛋白电泳、PCR扩增gyrA和parC基因、酶切分析和测序。结果：耐药株药物聚积量不及敏感株的1／2，经碳酰氰间氯苯腙（CCCP）处理后，聚积量上升并接近敏感株；经SDSPAGE电泳，耐药株与敏感株比较，外膜蛋白在约29ku条带处消失，而24ku处条带却明显增强；PCR－RFLP分析，gyrA基因能产生1条DNA片段，而parC基因则能产生2条片段；测序结果显示，其GyrA蛋白中所编码83位（相应于大肠埃希氏菌）氨基酸由丝氨酸变为亮氨酸，ParC蛋白未见氨基酸改变。结论：该株对环丙沙星耐药鲍氏不动杆菌存在DNA促旋酶变异，药物主动外运及外膜的改变。     

耐碳青霉烯类抗生素鲍曼不动杆菌耐药机制的研究

目的：调查和研究南昌地区鲍曼不动杆菌对碳青霉烯类抗生素的耐药机制。方法：收集南昌大学第二附属医院2012年间临床分离的耐亚胺培南鲍曼不动杆菌（CRAB）非重复株。Vitek-32型全自动微生物分析仪进行菌株鉴定,K-B法进行药敏试验,三维试验检测AmpC酶,EDTA协同试验检测金属β-内酰胺酶,采用聚合酶链反应（PCR）扩增耐药基因,并对阳性产物进行双向测序分析,确定其基因型。结果：84株CRAB对临床常用的12种抗菌药物有9种耐药率〉90%,对头孢哌酮-舒巴坦的耐药率最低（44.0%）,所有菌株均为多重耐药株。三维试验在83株菌（98.8%）中检出AmpC酶,协同试验未检出产金属酶菌株。所有菌株均携带blaOXA-23、blaOXA-51及blaADC基因,1株携带blaOXA-58基因,未检测到blaOXA-24及金属酶基因（blaIMP、blaVIM-2、blaNDM-1及blaSIM-1）;外排泵编码基因adeB、调控基因adeS和adeR的检出率分别为98.8%（83/84）、81.0%（68/84）和67.9%（57/84）;所有菌株均检出外膜蛋白carO基因;69株（82.1%）检测出I类整合酶基因（intI1）,全部菌株携带插入序列ISAba1。结论：南昌地区CRAB耐药性及多重耐药性非常严重,鲍曼不动杆菌对碳青霉烯类抗生素耐药的主要机制为产OXA-23型碳青霉烯酶,AmpC酶、外排泵AdeABC、I类整合子和插入序列ISAba1,可能与CRAB的多重耐药性密切相关。     

鲍曼不动杆菌的临床分离及其耐药性分析

目的 分析鲍曼不动杆菌（Ab）对抗菌药物的耐药性,以指导临床医师合理用药. 方法 应用法国生物-梅里埃ATB Expression型全自动鉴定等系统分析我院2009年1月1日至2010年2月28日住院患者分离出的56株Ab,对其分离率,耐药性进行分析. 结果 总分离率分别为：2009年1～4月1.25%（9/720）;2009年5～10月2.07%（15/726）;2009年11月至2010年2月4.34%（32/738）.其中重症监护室在各病区占的比例较高,百分比为53.6%,该菌在临床各种标本中的分布情况以百分比计其中痰标本高达87.5%. 结论 Ab目前对多黏菌素E无耐药,对亚胺培南、美罗培南敏感,对其他抗菌药物已高度耐药;重视细菌培养、合理使用抗菌药物有利于控制耐药菌的产生.     

Diversity of amino acid substitutions in PmrCAB associated with colistin resistance in clinical isolates of Acinetobacter baumannii

This study aimed to investigate the mechanisms of colistin resistance in 64 Acinetobacter baumannii isolates obtained from patients with ventilator-associated pneumonia hospitalised in Greece, Italy and Spain. In total, 31 A. baumannii isolates were colistin-resistant. Several novel amino acid substitutions in PmrCAB were found in 27 colistin-resistant A. baumannii. Most substitutions were detected in PmrB, indicating the importance of the histidine kinase for colistin resistance. In two colistin-resistant isolates, 93 amino acid changes were observed in PmrCAB compared with A. baumannii ACICU, and homologous recombination across different clonal lineages was suggested. Analysis of gene expression revealed increased pmrC expression in isolates harbouring pmrCAB mutations. Complementation of A. baumannii ATCC 19606 and ATCC 17978 with a pmrAB variant revealed increased pmrC expression but unchanged colistin MICs, indicating additional unknown factors associated with colistin resistance. Moreover, a combination of PmrB and PmrC alterations was associated with very high colistin MICs, suggesting accumulation of mutations as the mechanism for high-level resistance. The pmrC homologue eptA was detected in 29 colistin-susceptible and 26 colistin-resistant isolates. ISAba1 was found upstream of eptA in eight colistin-susceptible and one colistin-resistant isolate and eptA was disrupted by ISAba125 in two colistin-resistant isolates. Whilst in most isolates an association of eptA with colistin resistance was excluded, in one isolate an amino acid substitution in EptA (R127L) combined with a point mutation in ISAba1 upstream of eptA contributed to elevated colistin MICs. This study helps to gain an insight into the diversity and complexity of colistin resistance in A. baumannii.     

Exposure to phototoxic NSAIDs and quinolones is associated with an increased risk of melanoma

Purpose

Ultraviolet radiation exposure is the most important exogenous risk factor for cutaneous malignancies. It is possible that phototoxic drugs promote the development of cutaneous melanoma (CM) by intensifying the effect of ultraviolet light on the skin. We investigated the association between the use of common systemic phototoxic drugs and development of CM.

Methods

This study was a case–control study in a Dutch population-based cohort. The drug dispensing data was obtained from PHARMO, a Dutch drug dispensing and hospital admissions registry, and linked to PALGA, the nationwide pathology network of the Netherlands. The cases were patients diagnosed with pathologically confirmed primary CM between 1991 and 2004. Controls were sampled from the PHARMO population. Exposure to systemic phototoxic drugs was measured and included antimicrobial agents, diuretics, antipsychotic drugs, antidiabetic drugs, cardiac drugs, antimalarials and nonsteroidal anti-inflammatory drugs (NSAIDs). A multivariate conditional logistic regression analysis was performed to study the association between exposure to phototoxic drugs and CM.

Results

The study population included 1,318 cases and 6,786 controls. Any phototoxic drug during the study period was dispensed for 46 % of the cases and 43 % of the controls (p?=?0.012). The use of quinolones [odds ratio (OR) 1.33, 95 % confidence interval (CI) 1.01–1.76] and propionic acid derivative NSAIDs (OR 1.33, 95 % CI 1.14–1.54) had a positive association with CM.

Conclusions

Our study shows that the use of phototoxic drugs is associated with an increased risk of developing CM. Even a short-term use of phototoxic quinolones and propionic acid derivative NSAIDs may increase the risk for CM. Patient education to promote sun-protective behaviour is essential to avoid immediate adverse effects and possible long-term effects of phototoxic drugs.     

Rapid development of Acinetobacter baumannii resistance to tigecycline

A 53-year-old woman experienced a multidrug-resistant (MDR) Acinetobacter baumannii urinary tract infection 5 months after undergoing kidney and liver transplantation. The tigecycline minimum inhibitory concentration (MIC) for her A. baumannii isolate was 1.5 microg/ml; the patient received 2 weeks of therapy with intravenous tigecycline as a 100-mg loading dose followed by 50 mg every 12 hours, with no lapses in treatment and with resolution of the infection. Three weeks later, MDR A. baumannii was isolated from her sputum in the setting of clinical evidence of pneumonia, and tigecycline was restarted; the tigecycline MIC for the A. baumannii isolate was 2 microg/ml. At approximately the same time, the patient was found to have a paraspinal abscess and spinal osteomyelitis. Cultures of the abscess fluid grew A. baumannii with a tigecycline MIC of 24 microg/ml. A follow-up sputum culture again yielded A. baumannii, but with a tigecycline MIC of 24 microg/ml. Urine culture at that time also grew A. baumannii with a tigecycline MIC of 24 microg/ml. Clinicians should be aware that tigecycline MICs for A. baumannii isolates may increase during therapy with tigecycline after only brief exposure to the drug. Patients receiving tigecycline for Acinetobacter should be monitored for the development of clinical resistance, and isolates should be monitored for evidence of microbiologic resistance.     

Molecular epidemiology and mechanisms of rifampicin resistance in Acinetobacter baumannii isolates from Italy

Use of rifampicin (RIF) in combination with colistin (COL) has been proposed for the treatment of multidrug-resistant Acinetobacter baumannii infections owing to in vitro synergism. The aim of the present study was to evaluate the molecular epidemiology and mechanisms of RIF resistance in 57 clinical isolates of A. baumannii in two tertiary care hospitals in Naples (Italy) from 2006 to 2010. Amongst the collection, 36 isolates showed high RIF minimum inhibitory concentrations (MICs) (256 mg/L to ≥512 mg/L), 16 showed intermediate MICs (8-16 mg/L) and 5 had low MICs (4 mg/L). Of the 36 isolates with elevated RIF MICs, 35 were assigned to sequence type ST2 and 1 to ST78. Amongst the 57 isolates, 35 carried at least one mutation in rpoB, including H535L in 9 isolates and double mutations D525N and P544L in 7 isolates, whilst 22 showed no rpoB mutations. Treatment with the efflux pump inhibitor phenyl-arginine-β-naphthylamide (PAβN) of resistant isolates with no mutations in rpoB and different RIF MICs reduced the MIC by >10-fold and restored the synergism between RIF and COL in time-kill studies, whilst it had no effect on strains carrying rpoB mutations. In conclusion, the emergence of elevated RIF MICs in A. baumannii isolates from our geographical area was mostly caused by mutations in rpoB; low to intermediate RIF MICs were also caused by altered membrane permeability to the drug. The phenomenon was contributed by the selection of two prevalent clones both assigned to ST2 genotype. These data may have implications for the correct identification of cases with A. baumannii infection that would not benefit from addition of RIF to COL.     

临床分离鲍曼不动杆菌耐药性及qnr基因检测

目的了解临床分离鲍曼不动杆菌中qnr基因和ESBLs基因的分布及其耐药特征。方法采用PCR法对115株鲍曼不动杆菌进行qnrA、qnrB、qnrS基因筛查,并用PCR法检测qnr阳性菌株SHV、TEM、CTX-M-14及CTX-M-3型ESBLs基因;用琼脂对倍稀释法测定15种抗菌药物对qnr阳性株的MIC值。结果115株鲍曼不动杆菌中,2株(1.74%)细菌检出qnrB基因;qnr阳性菌株同时检出SHV、CTX-M-14、TEM、CTX-M-3型ESBLs基因。1株qnr阳性菌株对4种喹诺酮类耐药,1株对左氧氟沙星及莫西沙星中介,对环丙沙星和洛美沙星耐药;2株阳性菌除对亚胺培南和头孢哌酮/舒巴坦敏感外,对其他的β-内酰胺类抗菌药物均耐药。结论临床分离鲍曼不动杆菌中存在qnrB基因,qnr阳性株同时含有ESBLs基因,且呈多重耐药,临床应加强监测。     

Prevalence and analysis of microbiological factors associated with phenotypic heterogeneous resistance to carbapenems in Acinetobacter baumannii

The objectives of this study were to determine the prevalence of Acinetobacter baumannii with phenotypic heterogeneous resistance (PHR) to carbapenems (colonies inside the halo of inhibition) and to analyse its association with several microbiological variables. Acinetobacter baumannii isolates collected in Spain were used to analyse: (i) minimum inhibitory concentrations (MICs) of carbapenems; (ii) heteroresistance to carbapenems; (iii) genes encoding β-lactamases (bla genes); (iv) insertion sequences; and (v) inactivation of genes encoding porins (CarO, OprD and Omp33-36) and genes associated with the AdeABC efflux system (adeB, adeR and adeS). Polymerase chain reaction (PCR) amplification was used for gene detection. The rate of PHR was 20% to imipenem and 24% to meropenem. Susceptibility to imipenem was observed in 39% of PHR isolates. MICs of carbapenems for colonies were similar (± 1 log(2) dilution) to those of their parental isolates. These colonies growing inside the inhibition halo also reproduced the PHR to carbapenems. Differences observed between PHR isolates and non-PHR isolates were: bla(OXA-58-like), 57% vs. 0%; oprD-like, 96% vs. 56%; adeB, 89% vs. 94%; adeR, 82% vs. 94%; adeS, 82% vs. 94%; ISAba2, 61% vs. 31%; and ISAba3, 57% vs. 0%. No interruption of genes encoding porins or the efflux-related genes (adeB, adeR and adeS) was observed. In conclusion, A. baumannii strains with PHR to carbapenems are widespread in Spain. This phenotype is present in carbapenem-susceptible isolates as well as those that are not susceptible to carbapenems. Heteroresistance cannot explain the PHR to carbapenems, which appears to relate more to persistence or tolerance to carbapenems. bla(OXA-58-like), bla(OXA-51-like), ISAba2 and ISAba3 are associated with PHR to carbapenems. Inactivation of genes encoding porins or genes related to AdeABC is infrequent.     

鲍曼不动杆菌耐药表型与外排泵基因表达水平的研究

目的 探讨鲍曼不动杆菌临床分离株对常用抗菌药物的耐药性及与外排泵adeA基因表达水平之间的关系.方法 琼脂二倍稀释法检测鲍曼不动杆菌临床分离株对17种常用抗菌药物的最低抑菌浓度(MIC);PCR法扩增外排泵编码基因adeA:实时荧光定量RT-PCR(Real Time Fluorescent Quantitative RT-PCR)法检测adeA基因的mRNA表达水平.结果 药敏结果显示86株鲍曼不动杆菌对多粘菌素B全部敏感(100%),其次是美罗培南(73.2%)、亚胺培南(70.9%).耐药率最高的是庆大霉素(84.9%),其次为美罗西林(83.7%)和环丙沙星(79.1%).55株菌为多重耐药株(64%),其中5株(5/86)对多粘菌素B外的所有抗菌药物耐药(5.8%).adeA的检出阳性率为84.9%(73/86),Real Time RT-PCR结果显示多重耐药菌株adeA基因的mRNA相对表达量均高于敏感菌株,其中3株相对表达量为敏感菌株平均表达水平的30倍以上.结论 鲍曼不动杆菌临床分离株耐药情况严重,其主动外排系统adeA基因表达增强在多重耐药性形成中起重要作用.     

耐碳青霉烯鲍曼不动杆菌感染及预后的危险因素分析

目的：研究耐碳青霉烯类鲍氏不动杆菌（CRAB）感染的危险因素及死亡预后因素,提升临床对CRAB感染防控的认识。方法：采用病例对照性研究的方法,收集医院2016年1月至12月鲍曼不动杆菌感染患者112例,耐碳青霉烯类细菌（CRAB）感染组68例,碳青霉烯类抗菌药物敏感鲍曼不动杆菌（CSAB）感染组44例;应用单因素分析及多因素Logistic回归分析回顾性分析66例CRAB组危险因素及死亡危险因素。结果：单因素分析发现CRAB较CSAB的危险因素包括ICU入驻、同时分离多种致病菌、胃管、深静脉插管、尿管、机械通气（≥7 d）、碳青霉烯类抗菌药物。再进行Logistic多因素回归分析,结果显示机械通气和合并细菌感染是CRAB的独立危险因素。预后单因素分析,CRAB组死亡的危险因素包括：恶性肿瘤、ICU入驻、尿管、机械通气≥7 d、三代头孢/β内酰胺酶抑制剂。Logistic多因素回归分析发现恶性肿瘤是导致CRAB患者死亡的预后因素。结论：机械通气与合并细菌感染是CRAB发生的独立危险因素;恶性肿瘤是CRAB患者死亡的预后因素。     

鲍曼不动杆菌对主要抗菌药物耐药机制

近年来,鲍曼不动杆菌感染日益增多,并呈现多重耐药甚至是泛耐药趋势.本文就鲍曼不动杆菌对临床主要使用的抗菌药物的耐药机制研究进展做一综述.     

Epidemiological and clinical features associated with colonisation/infection by Acinetobacter baumannii with phenotypic heterogeneous resistance to carbapenems

The objective of this study was to identify risk factors for the acquisition of Acinetobacter baumannii with phenotypic heterogeneous resistance (PHR) to carbapenems and to determine whether these factors are similar to those associated with A. baumannii not showing this phenotype. Microbiological and clinical data from 211 patients included in the GEIH-Ab 2000 project were used. Isolates of A. baumannii were studied for their susceptibility to imipenem (IPM) by microdilution and for PHR to IPM as determined by the presence of colonies growing within the inhibition zone of IPM disks. Isolates were divided into three groups: (i) IPM-PHR isolates, i.e. susceptible and non-susceptible A. baumannii displaying PHR to IPM; (ii) non-IPM-PHR isolates, i.e. susceptible A. baumannii showing an inhibition halo but no colonies growing within it; and (iii) IPM-FR isolates, i.e. fully resistant A. baumannii displaying no halo of inhibition. IPM-PHR isolates of A. baumannii were more commonly isolated from respiratory tract samples and less commonly from urine, and were more frequently causes of infection than were IPM-FR isolates. Independent risk factors identified in patients with IPM-PHR isolates were Intensive Care Unit admission, surgery, and previous use of piperacillin/tazobactam or carbapenems, whilst risk factors for IPM-FR and IPM-PHR were previous use of cephalosporins and isolation from a urine sample. In conclusion, risk factors associated with colonisation/infection by isolates of A. baumannii with PHR to carbapenems are similar to those previously described for isolates resistant to carbapenems.     

Wide spread of Tn2006 in an AbaR4-type resistance island among carbapenem-resistant Acinetobacter baumannii clinical isolates in Taiwan

The bactericidal activity and resistance selectivity of garenoxacin against Streptococcus pneumoniae with mutations in ParC (S79F) or both GyrA (S81F) and ParC (D83Y and K137N) were investigated using in vitro pharmacokinetic models simulating plasma concentrations for a standard clinical regimen [400mg once daily (q.d.)]. The efficacy of garenoxacin was compared with that of levofloxacin (500 mg q.d.) and moxifloxacin (400mg q.d.). Garenoxacin showed excellent bactericidal activity against S. pneumoniae, including quinolone-resistant S. pneumoniae (QRSP), achieving ratios of area under the plasma concentration-time curve over 24h to minimum inhibitory concentration (AUC(0-24)/MIC) ≥ 26.3, without emerging resistant subpopulations. The area above the killing curves was greater and the time to achieve 99.9% killing was shorter for garenoxacin than the corresponding values for levofloxacin and moxifloxacin. No resistant subpopulations and no additional substitution of amino acids in GyrA or ParC emerged following treatment with garenoxacin. On the other hand, in the parC mutant strain, levofloxacin and moxifloxacin treatment caused an increase in the frequency of the resistant population and an additional substitution of amino acids in GyrA (levofloxacin, S81Y/F/C; moxifloxacin, S81Y or E85K). In QRSP with mutations in GyrA and ParC, levofloxacin had no bactericidal activity, whilst the bactericidal activity of moxifloxacin was less than that of garenoxacin; moreover, an additional substitution of amino acids in ParC (S79Y) was noted. In conclusion, garenoxacin corresponding to an oral dose of 400mg showed excellent bactericidal activity against S. pneumoniae, including QRSP, without the emergence of resistant mutants.     

耐碳青酶烯类鲍曼不动杆菌肺部感染患者adeB基因检测分析

目的:探讨耐碳青霉烯类鲍曼不动杆菌耐药性与adeB基因的关系,为临床治疗提供新思路.方法:收集2018年1月至2019年1月肺部感染患者痰液标本642份,分离鉴定鲍曼不动杆菌株,采用PCR检测其adeB基因表达,采用微量二倍稀释法检测MIC50.结果:分离出鲍曼不动杆菌250株,adeB基因表达异常的为152株.ade...     

耐亚胺培南鲍曼不动杆菌耐药性及OXA碳青霉烯酶检测

目的 分析97株临床分离的耐亚胺培南鲍曼不动杆菌的耐药特点及OxA碳青霉烯酶分布.方法 用国际标准平皿二倍稀释法,明确研究菌株的耐药表型;用脉冲场凝胶电泳法,对其进行分型并用PCR的方法,检测OXA碳青霉烯酶.结果 97株耐亚胺培南鲍曼不动杆菌均为多药耐药菌株,其中2株为泛耐药菌株;主要存在7个流行克隆株;有78株菌携带bal-oxa-23基因(80.4%);2株菌携带bal-oxa-58基因(2.1%);未发现携带bal-oxa-24基因的菌株.结论 在我国的亚胺培南耐药的鲍曼不动杆菌中,OXA-23仍为主要分布的碳青霉烯酶.     

Mechanisms of resistance to ciprofloxacin,ampicillin/sulbactam and imipenem in Acinetobacter baumannii clinical isolates in Taiwan

Nosocomial infections caused by multidrug-resistant (MDR) Acinetobacter baumannii have been increasing in recent years, posing a threat to public health worldwide. The susceptibility to eight antimicrobial agents of 35 clinical A. baumannii isolates from Taiwan was tested. Isolates were examined by polymerase chain reaction (PCR) and sequencing for β-lactamase genes and mutations in the gyrA and parC genes. Expression of AdeB, an efflux pump protein, was evaluated by real-time quantitative PCR. The level of adeB expression correlated with resistance to ciprofloxacin and ampicillin/sulbactam in A. baumannii isolates. Almost all isolates with full resistance to ciprofloxacin had both high adeB expression and point mutations in parC and gyrA, but 4 intermediate-resistant isolates had only high adeB expression without point mutations in gyrA or parC, in contrast to 18 susceptible isolates with low adeB expression and without mutations in gyrA or parC. Sixteen isolates (45.7%) carrying a type 1 integron were MDR as well as being more resistant to imipenem, amikacin, gentamicin, ceftazidime or cefepime than those without the integron. The class 1 integron in A. baumannii carried different resistance gene cassettes, including 5′CS-bla_IMP-1–aadA4–3′CS, 5′CS–aacA4–aadA1–3′CS and 5′CS–aacC1–aadA1–3′CS. In conclusion, expression of the adeB gene was associated with resistance to ciprofloxacin and ampicillin/sulbactam in A. baumannii. Multiple mutations in gyrA and parC also played a role in ciprofloxacin resistance. The major metallo-β-lactamase contributing to imipenem resistance in A. baumannii in Taiwan was bla_IMP-1, which was carried by the class 1 integron. The class 1 integron was associated with the MDR phenotype in A. baumannii.     

脂多糖缺失所致多黏菌素耐药的鲍曼不动杆菌研究进展

多黏菌素E(黏菌素)和多黏菌素B已成为治疗广泛耐药的革兰阴性菌感染的“最后一条防线”。但是，近年来有关鲍曼不动杆菌对多黏菌素的耐药性时有报道，尤其是有关脂多糖(LPS)缺失或修饰导致多黏菌素耐药的鲍曼不动杆菌各方面性质的改变，如细菌适应性下降，外膜合成相关基因表达增加，对其他抗生素敏感性提高，机械强度降低，细菌毒力下降，固有免疫因子水平变化等。本文将对近年来有关LPS缺失引起的鲍曼不动杆菌各方面性质变化的研究进行简要的综述。     

In vitro activity of doripenem against Acinetobacter baumannii clinical isolates

Doripenem is a carbapenem with activity against Gram-positive and Gram-negative pathogens. This study evaluated the in vitro activity of doripenem against a collection of 87 Acinetobacter baumannii clinical isolates, showing that the activity of doripenem was superior to imipenem and meropenem for strains carrying the bla(OXA-58) gene. A. baumannii clinical isolates expressing the bla(OXA-24) gene were resistant to doripenem, imipenem and meropenem. However, in clinical isolates expressing the bla(OXA-58) gene, the percentage of isolates with a doripenem minimum inhibitory concentration >8microg/mL was much lower than that of imipenem and meropenem. This study shows that the activity of doripenem was superior to imipenem and meropenem for strains carrying the bla(OXA-58) gene.