抗菌药物联合应用需求日益迫切，这4种体外联合药敏试验你是否都了解？|利福平|沙星|舒巴坦|菌株

*仅供医学专业人士参考阅读

进行抗菌药物组合需要体外联合药敏试验数据的支持。

近年来由于多重耐药细菌的日益增加以及治疗这类细菌感染的药物选择有限，特别是治疗由多重耐药菌引起的严重感染，因此，对新的抗菌药物或抗菌药物联合的临床应用需求变得越来越迫切。

中国医院内感染抗菌药物耐药监测（CARES）[1]回顾了2016年12家教学医院引起院内感染的2060例病例，碳青霉烯耐药的肺炎克雷伯菌和大肠埃希菌发生率分别为15.3%和1.8%，碳青霉烯耐药的铜绿假单胞菌发生率为31.7%。2015年中国CRE监测网[2]显示碳青霉烯类耐药肠杆菌科细菌（CRE）引起的死亡率为33.5%，其中血流感染43.1%，腹腔内感染26%，下呼吸道感染34.8%，尿路感染30.3%。而治疗多重耐药菌可选择的药物少，常需两药联合或三药联合[3,4]。

体外敏感率较高的药物仅有多黏菌素、替加环素等，既往曾认为多黏菌素、替加环素和部分氨基糖苷类可用于治疗，但是替加环素和多黏菌素由于药物代谢动力学/药效学（PK/PD）特性不尽人意，治疗效果也难以达到预期。多重耐药菌的治疗尚无理想的抗菌药物，很多情况下需要联合用药，如何进行抗菌药物组合需要体外联合药敏试验数据的支持。本文主要列举了几种体外联合药敏试验常用方法学，包括时间-杀菌曲线、棋盘稀释法、纸片法和基于Etest条的方法。

1.时间-杀菌曲线

时间-杀菌曲线（The time- kill assay，TKA）被认为是确定抗菌药物之间是否有协同作用的最有效方法。TKA是一种肉汤稀释的方法，需先测定拟组合抗菌药物对病原菌的最低抑菌浓度（MIC）值，再根据MIC值进行不同抗菌药物组合并对病原菌进行菌落计数。接种的菌量约为5x 105 CFU/mL，选择不同的时间间隔，一般选择的时间点为0、2、4、6、8、12、24、30、36、48小时，从含有抗菌药物的肉汤中进行传代培养，将菌液涂布在LB琼脂平板上，置于35˚C培养箱孵育后进行细菌菌落计数，最后菌落数以log 10 CFU/mL表示并绘制曲线图。对于浓度依赖性抗菌药物需在24h内每2h菌落计数一次，对于时间依赖性抗菌药物，可每3-4h进行一次菌落计数，直到24-48h。

TKA中协同作用的定义为与单一抗菌药物相比，组合抗菌药物中细菌生长量的减少≥2log10CFU/mL。拮抗作用的定义是与单一抗菌药物相比，组合中细菌生长量增加≥2log10CFU/mL。小于2log10CFU/mL的差异被解释为无相关性[5]。

另一种解释TKA的方法是杀菌曲线下面积（area under the killing curve， AUKC）[6] ，而不是计算细菌菌落数；在结果绘制的曲线图上，Y轴为 log10CFU/mL值，X轴为时间。任何P<0.05的统计学差异被视为协同相互作用。表1总结了采用时间杀菌曲线的方法对碳青霉烯类耐药的鲍曼不动杆菌、铜绿假单胞菌和肺炎克雷伯菌体外联合药敏试验的研究，对于鲍曼不动杆菌采用TKA的方法拟组合的抗菌药物主要是基于舒巴坦类和黏菌素的药物，铜绿假单胞菌拟组合的药物主要是碳青霉烯类和喹诺酮类药物，而对于肺炎克雷伯菌主要是基于替加环素、黏菌素/多黏菌素和碳青霉烯类药物，具体见表1。

表1 基于时间杀菌曲线对常见碳青霉烯类耐药菌株体外联合药敏试验的研究

Amp/Sul：氨苄西林/舒巴坦，Imi：亚胺培南，Col：黏菌素，Dori：多利培南，Tobra：妥布霉素，Sul：舒巴坦， Fos：磷霉素，Mero：美罗培南，PB：多黏菌素B，Rif：利福平，Tige：替加环素，Tmp：甲氧苄啶，Cotri：甲氧苄啶/磺胺甲恶唑，Cipro：环丙沙星，Gati：加替沙星，Cefe：头孢吡肟，Czd：头孢他啶，Gen：庆大霉素，Doxy：强力霉素，Ak：阿米卡星，ATM：氨曲南，CZA：头孢他啶/阿维巴坦

2.棋盘稀释法

棋盘稀释法（Checkerboard assay，CB）是用含有不同浓度抗菌药物组合的肉汤（2mL或100uL）在96孔板中操作的肉汤稀释法。分别测定拟组合的抗菌药物A和B对待测菌的MIC，根据抗菌药物A和B的MIC确定药物联合测定的稀释度，一般选择6～8个稀释度；每种抗菌药物的最高浓度为其MIC的2倍，置于35˚C培养箱孵育，判断抗菌药物组合对病原菌的抑制浓度范围。假设A药的MIC=32mg/L，B药的MIC=8mg/L，操作示意图见图1。结果判断采用FIC （Fractional inhibitory Concentration）指数[21]，FIC指数=A药联合时的MIC/A药单测时MIC+B药联合时的MIC/B药单测时MIC，判断标准：FIC指数≤0.5为协同作用；0.52为拮抗作用。表2总结了采用棋盘稀释法对碳青霉烯类耐药鲍曼不动杆菌、铜绿假单胞菌和肺炎克雷伯菌体外联合药敏试验的研究。

图1 棋盘稀释法操作示意图

表2 基于棋盘稀释法对常见碳青霉烯类耐药菌株体外联合药敏试验的研究

Mino: 米诺环素，Van：万古霉素，Pip/Tazo：哌拉西林/他唑巴坦，Levo：左氧氟沙星，Genta：庆大霉素

3.基于Etest条的方法

含有抗菌药物浓度的Etest条已被确定可用于体外联合药敏试验方法，主要的方法有：1）Etest条交叉法，2）Etest条重叠法，3）Etest条琼脂法。

3.1 Etest条交叉法

在涂布待测菌的MH琼脂平板上分别用拟组合测试的抗菌药物Etest条测定单个抗菌药物的MIC值，置于35˚C培养箱培养并记录MIC值，再将2个拟组合抗菌药物Etest条90˚交叉置于在含有0.5麦氏浊度待测菌的MH琼脂平板上，两条Etest交叉点为单药测定是的MIC值位置，置于35˚C培养箱培养18h，记录抗菌药物组合时的MIC值并计算FIC指数[38]。例如：单药A的MIC=6，单药B的MIC=12，A联合时MIC=0.125，B联合时MIC=0.5，FIC指数=0.125/6+0.5/12=0.0625<0.5，因此抗菌药物A和B有协同作用，示意图见图2。

图2 Etest条交叉法示意图

3.2 Etest条重叠法

在这种方法中，MHA琼脂平板接种了0.5 麦氏浊度的待测菌。放置第一种含有抗菌药物的Etest条并在室温下孵育1小时，让抗菌药物扩散到培养基中。1 小时后，将其移除并记录初始MIC。然后将第二种含有抗菌药物的Etest条直接放在第一个试纸的印记上，置于35˚C培养箱培养18h，记录抗菌药物组合时的MIC值并计算FIC指数[39,40]。单药A的MIC=6，单药B的MIC=8，A+B联合时MIC=1.5，FIC指数=1.5/6+1.5/8=0.4375<0.5，因此抗菌药物A和B有协同作用，示意图见图3。

图3 Etest条重叠法示意图

3.3 Etest条琼脂法

在这种方法中[40]，先测定单药A的MIC，再使用0.5/0.125× MIC 的A药与琼脂粉的混合溶液制备MH平板，并将单药B的E test条放置在该琼脂平板上，孵育后的抑菌圈即为两者的组合MIC值，反之亦如此。当含有抗菌药物培养基的 MIC 降低三倍以上时，就说明两种抗菌药物存在协同作用。例如单药A的MIC=6，B联合时MIC=0.75，单药B的MIC=64，A联合时MIC=0.19，A和B联合时的MIC降低原来的三倍以上，因此A和B有协同作用，示意图见图4。

图4 Etest条琼脂法示意图

表3总结了采用Etest条的方法对碳青霉烯类耐药的鲍曼不动杆菌、铜绿假单胞菌和肺炎克雷伯菌体外联合药敏试验的研究，对于鲍曼不动杆菌、铜绿假单胞菌和肺炎克雷伯菌的协同率分别为0%-100%，3%-73.3%和0%-100%，具体的拟组合抗菌药物及其协同率见表3。

表3 基于Etest条的方法对常见碳青霉烯类耐药菌株体外联合药敏试验的研究

Netil：奈替米星，Tec：替考拉宁，ColR：黏菌素耐药

4.纸片法

4.1 滤纸条浸泡法

在该方法中，将浸泡在拟组合抗菌药物MIC或MIC以上溶液中的滤纸条以直角放置在接种在含有待测菌的MH琼脂平板上。滤纸条中的抗菌药物可以扩散在培养基中，在37°C下孵育18-24小时后观察抑菌圈生长方式，抑菌圈生长方式的解释如下图5：(a)协同作用（交界处抑菌圈扩宽）；(b)相加作用（交界处出现抑菌圈）；(c)拮抗作用（交界处出现抑菌圈缩小）；(d)无相关作用（抑菌圈无影响）。该方法首次由美国学者Victor Lorian 和Georgeta Fodor提出[51]，证实了甲氧苄氨嘧啶和磺胺甲恶唑对85%的大肠埃希菌、86%的克雷伯菌属和89%的奇异变形杆菌有协同作用，但该方法只提供了定性的结果，并未得到广泛的应用评价。

图5 滤纸条浸泡法

4.2 双纸片协同试验

双纸片协同试验通常用于检测超广谱β-内酰胺酶（ESBL）的产生，也可用于检测抗菌药物组合之间的协同作用。该方法在1997年I.M.Gould和K.Milne提出[52]，测定了哌拉西林/他唑巴坦与庆大霉素和环丙沙星在屎肠球菌、金黄色葡萄球菌、表皮葡萄球菌、化脓性链球菌、嗜麦芽窄食单胞菌、铜绿假单胞菌、柠檬酸杆菌属、沙雷菌属、不动杆菌属和肠杆菌属之间的协同作用。在这种方法中，在涂布待测菌的MH琼脂平板上，拟组合的抗菌药物纸片的中心点距离约20mm，并在37˚C孵育。与单一抗菌药物的抑菌圈相比，直径增加≥2mm则表示有协同作用（6a），直径增加≤2mm则被归类为弱协同作用，若在两种抗菌药物之间交界处出现抑菌圈则为有相加作用（6b），而在两种抗菌药物抑菌圈连接处出现截断面为拮抗作用（6c），若两种抗菌药物抑菌圈直径相对于单药无变化则表明没有相关作用（6d），见图6。

图6 双纸片协同试验示意图

尽管该方法操作简单，结果易判断，但由于只能定性和存在主观解释，尚未得到广泛应用[53,54]。因此有研究认为双纸片协同试验适合用于铜绿假单胞菌的体外协同试验，对于铜绿假单胞菌来说，该方法比棋盘稀释法与临床疗效相比表现出更高的一致性[53]。表4总结了不同体外联合药敏试验方法的相对优缺点。

表4 不同体外联合药敏试验方法的相对优缺点

体外协同研究的结果转化为临床疗效的可能性仍然存在争议。有研究表明[55,56]，对于中性粒细胞减少或非中性粒细胞减少患者中的革兰氏阴性菌感染，β-内酰胺和氨基糖苷联合治疗没有临床益处，联合治疗可能有肾毒性的副作用。尽管存在毒性问题，但很多情况下联合治疗仍然是治疗多重耐药菌感染的最优选择。Biancofiore等人[57]报告了一名16岁女性多重耐药鲍曼不动杆菌的多灶性感染，在通过CB证明体外黏菌素、利福平和美罗培南的协同作用后，临床联合使用黏菌素、利福平和美罗培南治疗成功。Lee等人[58]报告了四名由鲍曼不动杆菌引起感染的患者（两名VAP患者和两名导管相关血流感染患者）使用碳青霉烯类和舒巴坦钠联合治疗的良好结局。通过CB分析对4株鲍曼不动杆菌进行的体外协同试验表明，FIC指数在0.56至0.75之间，舒巴坦钠和美罗培南/亚胺培南联合使用时有部分协同作用。Nastro等人[49]报告了黏菌素和利福平联合治疗由碳青霉烯类耐药肠杆菌引起的1例败血症、1例脑膜炎和1例复杂性尿路感染的成功案例，通过Etest琼脂法发现以上两种抗菌药物对所有患者的分离菌株都有协同作用。表5总结了体外协同药敏试验与临床治疗结局相关性的病例报道研究。

表5 体外协同药敏试验与临床治疗结局相关性的病例报道

对于减缓耐药菌的增长以及降低治疗耐药菌药物的毒副作用，联合治疗越来越多的受到关注。为给予临床在抗菌药物联合治疗上提供数据，体外联合药敏试验在两种或两种以上的抗菌药物的联合使用提供了支持。但该类试验方法的研究报道比较少，关于这些药物组合的协同、相加、无关、拮抗的数据也有限。虽然有许多试验方法可以确定抗菌药物之间的相互作用，但它们并没有很好地标准化，对测试结果所遵循的解释标准定义仍然不确定。本综述列举有几种方法可用于评估两种或多种抗菌药物的协同活性。然而，在技术问题、复杂性和测试结果的解释方面观察到了很大的差异。目前，TKA和CB作为参考方法，在各种研究中有较高一致率。但是，大多数体外试验方法不能预测临床成功率。因此，需要体外协同试验数据的前瞻性临床试验来指导用药改善临床结局，这意味着需要在全球范围内对各种方法进行标准化，以确定抗菌药物组合的协同作用。

参考文献：

1.李荷楠, et al. 2016年中国12家教学医院院内感染常见病原菌的分布和抗菌药物耐药监测研究. 中华检验医学杂志, 2018. 41(09): p. 651-657.

2.Zhang, Y, et al. Epidemiology of Carbapenem-Resistant Enterobacteriaceae Infections: Report from the China CRE Network. Antimicrob Agents Chemother, 2018. 62(2).

3.Yin, D, et al. Results from the China Antimicrobial Surveillance Network (CHINET) in 2017 of the In Vitro Activities of Ceftazidime-Avibactam and Ceftolozane-Tazobactam against Clinical Isolates of Enterobacteriaceae and Pseudomonas aeruginosa. Antimicrob Agents Chemother, 2019. 63(4).

4.Guan, X, et al. Laboratory diagnosis, clinical management and infection control of the infections caused by extensively drug-resistant Gram-negative bacilli: a Chinese consensus statement. Clin Microbiol Infect, 2016. 22 Suppl 1: p. S15-25.

5.Ko, W.C., et al. In vitro and in vivo combinations of cefotaxime and minocycline against Aeromonas hydrophila. Antimicrob Agents Chemother, 2001. 45(4): p. 1281-3.

6.MacGowan, A.P., et al. A new time-kill method of assessing the relative efficacy of antimicrobial agents alone and in combination developed using a representative beta-lactam, aminoglycoside and fluoroquinolone. J Antimicrob Chemother, 1996. 38(2): p. 193-203.

7.Sheng, W.H., et al. Comparative in vitro antimicrobial susceptibilities and synergistic activities of antimicrobial combinations against carbapenem-resistant Acinetobacter species: Acinetobacter baumannii versus Acinetobacter genospecies 3 and 13TU. Diagn Microbiol Infect Dis, 2011. 70(3): p. 380-6.

8.Principe, L., et al. In vitro activity of doripenem in combination with various antimicrobials against multidrug-resistant Acinetobacter baumannii: possible options for the treatment of complicated infection. Microb Drug Resist, 2013. 19(5): p. 407-14.

9.Choi, J.Y., et al. Synergic in-vitro activity of imipenem and sulbactam against Acinetobacter baumannii. Clin Microbiol Infect, 2004. 10(12): p. 1098-101.

10.Santimaleeworagun, W., et al. In vitro activity of colistin or sulbactam in combination with fosfomycin or imipenem against clinical isolates of carbapenem-resistant Acinetobacter baumannii producing OXA-23 carbapenemases. Southeast Asian J Trop Med Public Health, 2011. 42(4): p. 890-900.

11.Laishram, S., et al. Determination of synergy between sulbactam, meropenem and colistin in carbapenem-resistant Klebsiella pneumoniae and Acinetobacter baumannii isolates and correlation with the molecular mechanism of resistance. J Chemother, 2016. 28(4): p. 297-303.

12.Tan, T.Y., et al. In vitro antibiotic synergy in extensively drug-resistant Acinetobacter baumannii: the effect of testing by time-kill, checkerboard, and Etest methods. Antimicrob Agents Chemother, 2011. 55(1): p. 436-8.

13.Pankuch, G.A., et al. Activity of meropenem with and without ciprofloxacin and colistin against Pseudomonas aeruginosa and Acinetobacter baumannii. Antimicrob Agents Chemother, 2008. 52(1): p. 333-6.

14.Pankey, G.A. and D.S. Ashcraft, In vitro synergy of ciprofloxacin and gatifloxacin against ciprofloxacin-resistant Pseudomonas aeruginosa. Antimicrob Agents Chemother, 2005. 49(7): p. 2959-64.

15.Giamarellos-Bourboulis, E.J., et al. In vitro interaction of colistin and rifampin on multidrug-resistant Pseudomonas aeruginosa. J Chemother, 2003. 15(3): p. 235-8.

16.Pankey, G.A., D.S. Ashcraft, and A. Dornelles, Comparison of 3 Etest(®) methods and time-kill assay for determination of antimicrobial synergy against carbapenemase-producing Klebsiella species. Diagn Microbiol Infect Dis, 2013. 77(3): p. 220-6.

17.Jernigan, M.G., et al. The combination of doripenem and colistin is bactericidal and synergistic against colistin-resistant, carbapenemase-producing Klebsiella pneumoniae. Antimicrob Agents Chemother, 2012. 56(6): p. 3395-8.

18.Yim, H., et al. Time-kill synergy tests of tigecycline combined with imipenem, amikacin, and ciprofloxacin against clinical isolates of multidrug-resistant Klebsiella pneumoniae and Escherichia coli. Ann Clin Lab Sci, 2011. 41(1): p. 39-43.

19.Souli, M., et al. Does the activity of the combination of imipenem and colistin in vitro exceed the problem of resistance in metallo-beta-lactamase-producing Klebsiella pneumoniae isolates? Antimicrob Agents Chemother, 2009. 53(5): p. 2133-5.

20.Emeraud, C., et al. Aztreonam plus Clavulanate, Tazobactam, or Avibactam for Treatment of Infections Caused by Metallo-β-Lactamase-Producing Gram-Negative Bacteria. Antimicrob Agents Chemother, 2019. 63(5).

21.EUCAST Definitive Document E.Def 1.2, May 2000: Terminology relating to methods for the determination of susceptibility of bacteria to antimicrobial agents. Clin Microbiol Infect, 2000. 6(9): p. 503-8.

22.Hornsey, M., et al. In vitro activity of the novel monosulfactam BAL30072 alone and in combination with meropenem versus a diverse collection of important Gram-negative pathogens. Int J Antimicrob Agents, 2013. 42(4): p. 343-6.

23.Kiffer, C.R., et al. In vitro synergy test of meropenem and sulbactam against clinical isolates of Acinetobacter baumannii. Diagn Microbiol Infect Dis, 2005. 52(4): p. 317-22.

24.Ji, J., et al. In vitro activity of sulbactam in combination with imipenem, meropenem, panipenem or cefoperazone against clinical isolates of Acinetobacter baumannii. Int J Antimicrob Agents, 2013. 41(4): p. 400-1.

25.Pongpech, P., et al. Antibacterial activity of carbapenem-based combinations againts multidrug-resistant Acinetobacter baumannii. J Med Assoc Thai, 2010. 93(2): p. 161-71.

26.Ni, W., et al. In vitro effects of tigecycline in combination with colistin (polymyxin E) and sulbactam against multidrug-resistant Acinetobacter baumannii. J Antibiot (Tokyo), 2013. 66(12): p. 705-8.

27.Pei, G., Y. Mao, and Y. Sun, In vitro activity of minocycline alone and in combination with cefoperazone-sulbactam against carbapenem-resistant Acinetobacter baumannii. Microb Drug Resist, 2012. 18(6): p. 574-7.

28.Tong, W., et al. In vitro activity of cefepime combined with sulbactam against clinical isolates of carbapenem-resistant Acinetobacter spp. Int J Antimicrob Agents, 2006. 28(5): p. 454-6.

29.Mitsugui, C.S., et al. In vitro activity of polymyxins in combination with β-lactams against clinical strains of Pseudomonas aeruginosa. Int J Antimicrob Agents, 2011. 38(5): p. 447-50.

30.Piccoli, L., et al. In vitro and in vivo synergy of levofloxacin or amikacin both in combination with ceftazidime against clinical isolates of Pseudomonas aeruginosa. J Chemother, 2005. 17(4): p. 355-60.

31.Dawis, M.A., et al. In vitro activity of gatifloxacin alone and in combination with cefepime, meropenem, piperacillin and gentamicin against multidrug-resistant organisms. J Antimicrob Chemother, 2003. 51(5): p. 1203-11.

32.Visalli, M.A., M.R. Jacobs, and P.C. Appelbaum, Determination of activities of levofloxacin, alone and combined with gentamicin, ceftazidime, cefpirome, and meropenem, against 124 strains of Pseudomonas aeruginosa by checkerboard and time-kill methodology. Antimicrob Agents Chemother, 1998. 42(4): p. 953-5.

33.Tascini, C., et al. Synergistic activity of colistin plus rifampin against colistin-resistant KPC-producing Klebsiella pneumoniae. Antimicrob Agents Chemother, 2013. 57(8): p. 3990-3.

34.Clock, S.A., et al. In vitro activity of doripenem alone and in multi-agent combinations against extensively drug-resistant Acinetobacter baumannii and Klebsiella pneumoniae. Diagn Microbiol Infect Dis, 2013. 76(3): p. 343-6.

35.Elemam, A., J. Rahimian, and M. Doymaz, In vitro evaluation of antibiotic synergy for polymyxin B-resistant carbapenemase-producing Klebsiella pneumoniae. J Clin Microbiol, 2010. 48(10): p. 3558-62.

36.Gaibani, P., et al. In vitro activity and post-antibiotic effects of colistin in combination with other antimicrobials against colistin-resistant KPC-producing Klebsiella pneumoniae bloodstream isolates. J Antimicrob Chemother, 2014. 69(7): p. 1856-65.

37.Stein, C., et al. Three Dimensional Checkerboard Synergy Analysis of Colistin, Meropenem, Tigecycline against Multidrug-Resistant Clinical Klebsiella pneumonia Isolates. PLoS One, 2015. 10(6): p. e0126479.

38.White, R.L., et al. Comparison of three different in vitro methods of detecting synergy: time-kill, checkerboard, and E test. Antimicrob Agents Chemother, 1996. 40(8): p. 1914-8.

39.Manno, G., et al. Use of the E test to assess synergy of antibiotic combinations against isolates of Burkholderia cepacia-complex from patients with cystic fibrosis. Eur J Clin Microbiol Infect Dis, 2003. 22(1): p. 28-34.

40.Sopirala, M.M., et al. Synergy testing by Etest, microdilution checkerboard, and time-kill methods for pan-drug-resistant Acinetobacter baumannii. Antimicrob Agents Chemother, 2010. 54(11): p. 4678-83.

41.Cıkman, A., et al. [Evaluation of Colistin-Ampicillin/Sulbactam Combination Efficacy in Imipenem-Resistant Acinetobacter baumannii Strains]. Mikrobiyol Bul, 2013. 47(1): p. 147-51.

42.Kiratisin, P., A. Apisarnthanarak, and S. Kaewdaeng, Synergistic activities between carbapenems and other antimicrobial agents against Acinetobacter baumannii including multidrug-resistant and extensively drug-resistant isolates. Int J Antimicrob Agents, 2010. 36(3): p. 243-6.

43.Marie, M.A., et al. A prospective evaluation of synergistic effect of sulbactam and tazobactam combination with meropenem or colistin against multidrug resistant Acinetobacter baumannii. Bosn J Basic Med Sci, 2015. 15(4): p. 24-9.

44.Tan, T.Y., et al. In vitro effect of minocycline and colistin combinations on imipenem-resistant Acinetobacter baumannii clinical isolates. J Antimicrob Chemother, 2007. 60(2): p. 421-3.

45.Miyasaki, Y., et al. In vitro activity of antibiotic combinations against multidrug-resistant strains of Acinetobacter baumannii and the effects of their antibiotic resistance determinants. FEMS Microbiol Lett, 2012. 328(1): p. 26-31.

46.He, W., et al. In vitro Etest synergy of doripenem with amikacin, colistin, and levofloxacin against Pseudomonas aeruginosa with defined carbapenem resistance mechanisms as determined by the Etest method. Diagn Microbiol Infect Dis, 2012. 74(4): p. 417-9.

47.Samonis, G., et al. Synergy of fosfomycin with carbapenems, colistin, netilmicin, and tigecycline against multidrug-resistant Klebsiella pneumoniae, Escherichia coli, and Pseudomonas aeruginosa clinical isolates. Eur J Clin Microbiol Infect Dis, 2012. 31(5): p. 695-701.

48.Balke, B., et al. Evaluation of the E test for the assessment of synergy of antibiotic combinations against multiresistant Pseudomonas aeruginosa isolates from cystic fibrosis patients. Eur J Clin Microbiol Infect Dis, 2006. 25(1): p. 25-30.

49.Nastro, M., et al. Activity of the colistin-rifampicin combination against colistin-resistant, carbapenemase-producing Gram-negative bacteria. J Chemother, 2014. 26(4): p. 211-6.

50.Pankey, G.A. and D.S. Ashcraft, Detection of synergy using the combination of polymyxin B with either meropenem or rifampin against carbapenemase-producing Klebsiella pneumoniae. Diagn Microbiol Infect Dis, 2011. 70(4): p. 561-4.

51.Lorian, V. and G. Fodor, Technique for determining the bactericidal effect of drug combinations. Antimicrob Agents Chemother, 1974. 5(6): p. 630-3.

52.Gould, I.M. and K. Milne, In-vitro pharmacodynamic studies of piperacillin/tazobactam with gentamicin and ciprofloxacin. J Antimicrob Chemother, 1997. 39(1): p. 53-61.

53.Mayer, I. and E. Nagy, Investigation of the synergic effects of aminoglycoside-fluoroquinolone and third-generation cephalosporin combinations against clinical isolates of Pseudomonas spp. J Antimicrob Chemother, 1999. 43(5): p. 651-7.

54.Altoparlak, U., et al. Prevalence of metallo-beta-lactamase among Pseudomonas aeruginosa and Acinetobacter baumannii isolated from burn wounds and in vitro activities of antibiotic combinations against these isolates. Burns, 2005. 31(6): p. 707-10.

55.Marcus, R., et al. Clinical implications of β-lactam-aminoglycoside synergism: systematic review of randomised trials. Int J Antimicrob Agents, 2011. 37(6): p. 491-503.

56.Paul, M., et al. Beta-lactam versus beta-lactam-aminoglycoside combination therapy in cancer patients with neutropenia. Cochrane Database Syst Rev, 2013. 2013(6): p. Cd003038.

57.Biancofiore, G., et al. Colistin, meropenem and rifampin in a combination therapy for multi-drug-resistant Acinetobacter baumannii multifocal infection. A case report. Minerva Anestesiol, 2007. 73(3): p. 181-5.

58.Lee, N.Y., et al. Combination carbapenem-sulbactam therapy for critically ill patients with multidrug-resistant Acinetobacter baumannii bacteremia: four case reports and an in vitro combination synergy study. Pharmacotherapy, 2007. 27(11): p. 1506-11.

59.Lee, C.H., et al. Antimicrobial effects of varied combinations of meropenem, sulbactam, and colistin on a multidrug-resistant Acinetobacter baumannii isolate that caused meningitis and bacteremia. Microb Drug Resist, 2008. 14(3): p. 233-7.

专家简介

康梅

主任技师，医学博士，硕士研究生导师

四川大学华西医院实验医学科临床微生物室