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

Treatment of infections with carbapenem-resistant Gram negative organism is a major challenge especially among intensive care patients. Combinations of sulbactam, meropenem and colistin was studied for its synergistic activity against 100 invasive isolates of carbapenem-resistant Klebsiella pneumoniae and Acinetobacter baumannii-calcoaceticus complex by checkerboard assay and time kill assay (TKA). In addition, presence of carbapenemase production was determined by multiplex PCR. Time kill assay detected more synergy than checkerboard assay. Good bactericidal activity of 70–100% was noted with the combinations tested. Among K. pneumoniae, isolates producing NDM carbapenemase alone showed significantly more synergy than isolates producing OXA-48-like carbapenemases. In treatment of infection with carbapenem-resistant organisms, the site of infection and the type of carbapenemase produced may help to determine the most effective combination of antimicrobials.     

Trends in the resistance profiles of Acinetobacter baumannii endemic clones in a university hospital of Argentina

A total of 925 Acinetobacter spp. isolates were collected from routine clinical samples of patients admitted to the university hospital of Buenos Aires city during the period 2004–2012. From this collection, 129 isolates identified as Acinetobacter baumannii were selected for molecular studies. Minimal inhibitory concentrations (MICs) of antimicrobials were determined by agar dilution method. Colistin (COL) heteroresistance was investigated by means of population analysis studies. PCR-based methods were used for epidemiological analysis and for the screening of carbapenemases and the bla_tetB gene. We have observed a steady rise in the MIC₅₀ of imipenem (IMI)-resistant isolates and an increment in the presence of bla_OXA-23-like gene (74–100%) as well. A rapid increasing rate of minocycline (MIN) resistance and a rise of the MIC₅₀ of the resistant isolates have been detected since the year 2008. All isolates harboured the tet (B) gene. An increase in the value of the tigecycline (TIG) MIC was seen from the year 2007 onwards. This loss of activity was observed among different clones. A rise of COL heteroresistance from 46.4% in 2004 to 95% in 2012 was detected. During this period, COL consumption also increased (11.1-fold). However, COL resistance remained sporadic.     

A systematic review of implications,mechanisms, and stability of in vivo emergent resistance to colistin and tigecycline in Acinetobacter baumannii

The potential of A. baumannii for acquired resistance to last resort antibiotics (colistin and tigecycline) during treatment has important clinical implications, especially when dealing with patients failing to improve despite treatment with an active antimicrobial. However, the relevant literature remains scattered. Therefore, a systematic search was conducted in PubMed and Scopus. Several studies reported emergence of resistance to colistin or tigecycline during treatment, in most cases (86%) resulting in persistent or recurrent infections, especially in cases of emergent resistance without fitness cost. Lipopolysaccharide modification in the case of colistin and overexpression of efflux pumps in the case of tigecycline were the main mechanisms of resistance. Emergent colistin resistance is often associated with fitness cost which may result in re-emergence of the fitter and more virulent colistin susceptible strain after cessation of antibiotic pressure. Prospective studies are needed to determine the frequency of emergent resistance during treatment and its impact on patient outcomes.     

In Vitro Interactions of Aminoglycosides with Imipenem or Ciprofloxacin against Aminoglycoside Resistant Acinetobacter baumannii

Summary

The in vitro interactions between gentamicin, tobramycin, netilmicin and amikacin with imipenem and ciprofloxacin were evaluated by the killing curve technique against 20 clinical isolates of Acinetobacter baumannii highly resistant to aminoglycosides which were susceptible or moderately susceptible to imipenem and resistant or moderately susceptible to ciprofloxacin. Imipenem enhanced killing by gentamicin, tobramycin, netilmicin and amikacin in tests with 9, 12, 10 and 15 strains (45-75%) while ciprofloxacin with 3, 7, 5 and 6 strains (15-35%) respectively. Interaction results were influenced by the height of aminoglycoside minimum bactericidal concentrations (MBCs) but were independent of imipenem or ciprofloxacin MBCs and the presence of aminoglycoside modifying enzymes. It is concluded that enhanced killing after aminoglycoside interaction with imipenem or ciprofloxacin versus A. baumannii cannot be predicted but it should be carefully tested in vitro. The in vivo significance of the reported findings mandates clinical studies in humans.     

Antibacterial and anti-biofilm activities of melittin and colistin,alone and in combination with antibiotics against Gram-negative bacteria

In vitro antibacterial and anti-biofilm activities of antimicrobial cationic peptides (AMPs) – melittin and colistin – both alone and in combination with antibiotics were evaluated against clinical isolates of Gram-negative bacteria. Minimum inhibitory concentration (MIC) and fractional inhibitory concentration (FIC) index were determined by the microbroth dilution and chequerboard techniques, respectively. The time-kill curve (TKC) method was used for determining the bactericidal activities of AMPs alone and in combination. Measurements of anti-biofilm activities were performed spectrophotometrically for both inhibition of attachment and 24-hour biofilm formation at MIC or subMIC. According to MIC₉₀ values, the most active agents against Pseudomonas aeruginosa, Escherichia coli and Klebsiella pneumoniae were colistin, imipenem and ciprofloxacin, respectively. In combination studies, synergistic effects were mostly seen with colistin–imipenem against E. coli and K. pneumoniae (50 and 54%, respectively), colistin–ciprofloxacin against P. aeruginosa (77%). In TKC studies, synergism was observed with almost all expected combinations, even more frequently than chequerboard method. All of the antimicrobial agents were able to inhibit attachment and 24-hour biofilm formation between 0–57% at 1/10?×?MIC and 7–73% at 1?×?or 1/10?×?MIC, respectively. AMPs seem to be a good candidate for antimicrobial chemotherapy with their antibacterial and anti-biofilm activities as a single agent or in combination with antibiotics.     

The resistance and transmission mechanism of Acinetobacter baumannii isolates in a tertiary care hospital,China

The aim of this study was to analyse the resistance and epidemiological data of 117 Acinetobacter baumannii isolates from Southwest Hospital, Chongqing, China. Except for polymyxin B, tigecycline, minocycline, cefoperazone/sulbactam, amikacin and levofloxacin, the resistance rates of other antimicrobial agents were above 90%. All the clinical isolates had the bla_OXA-51 gene and 114 isolates had the bla_OXA-23 gene. Forty-nine isolates were found to contain the bla_IMP-4 gene. PFGE data showed that 117 isolates were divided into 25 groups. Sixty-three (53.85%) were found to carry the class 1 integron, and the sequence analysis of the class 1 integron internal variable regions – five types, one of which had the bla_IMP-4 gene. For the bla_IMP-4 positive strain without class 1 integron, we found the flanking sequence had the TnpA gene. The result suggested that the resistance gene was widely distributed in our hospital; moreover, the modes of presence and transmission are different and complicated. The results of our study can improve the infection empirical treatment method and infection control programme.     

Characterization of carbapenem resistance mechanisms in Klebsiella pneumoniae and in vitro synergy of the colistin–meropenem combination

Abstract

In this prospective study, consecutive isolates of Klebsiella pneumoniae were tested for different mechanisms of carbapenem resistance using the modified Hodge test (MHT), Rosco Neo-Sensitabs (ROSCO). Phenylalanine arginine beta-naphthylamide assay (PABN) inhibitor-based test was done on isolates in which the mechanism of resistance was not identifiable by the ROSCO. Among 105 selected isolates, carbapenemase production was noted in 100 (95%) by MHT and ROSCO showed 97 (92·4%) inhibition with dipicolinic acid signifying the production of MBL. PCR amplification was positive in 90 (86%) isolates for bla_NDM-1 and 46 (44%) isolates for bla_OXA-48. 54 (51%) isolates were positive for bla_CTX-M and all belonged to bla_CTX-M group 1. Isolates co produced bla_OXA-48 (31/105, 30%) and bla_CTX-M (40/105, 38%) in combination with the carbapenemase (bla_NDM-1) gene. Five colistin-resistant isolates were positive for bla_OXA-48. Eight isolates did not show inhibition with any of the inhibitor containing disks and found to be positive for bla_OXA-48. Isolates were tested for colistin-meropenem synergy and detection rate was higher by the checkerboard (48%) than E-test method (35%). Our study necessitates continuous surveillance to recognize the predominant machinery of resistance in a particular geographical region to formulate effective control measures.     

In Vitro Evaluation of the Antimicrobial Activity of Meropenem in Combination with Polymyxin B and Gatifloxacin Against Pseudomonas aeruginosa and Acinetobacter baumannii

Abstract

The antimicrobial activity of meropenem combined with either polymyxin B or gatifloxacin was evaluated by the checkerboard method against Pseudomonas aeruginosa (10 strains) and Acinetobacter baumannii (10 strains). In addition, the triple combination of polymyxin B, gatifloxacin, and meropenem was also studied as well as the polymyxin B and gatifloxacin combination. A partial synergism interaction between meropenem and polymyxin B was observed for 80% of the A. baumannii strains. In contrast, this combination showed an indifferent effect for 80% of the P. aeruginosa strains tested. The combination of meropenem and gatifloxacin showed synergism only for two strains of A. baumannii, and partial synergism and additive effect for seven strains and indifference for four strains of both species. For the strains of P. aeruginosa, the double combination of polymyxin B and gatifloxacin and the triple combination of meropenem, polymyxin B and gatifloxacin were indifferent for the majority of the strains tested, that is, 90 and 80% respectively.     

Antitumor effect of lonidamine alone or combined with tamoxifen or medroxyprogesterone acetate in breast cancer cells.

The effect of Lonidamine (LND) alone or combined with the antiestrogen Tamoxifen (TAM) or Medroxyprogesterone acetate (MPA) on cell proliferation and steroid hormone receptor content of a human estrogen sensitive breast cancer cell line was investigated. LND has a direct growth inhibitory action, even if used at relatively low concentrations (10(-7) M), and shows the maximum effect at 10(-4) M. The combination of LND with the antiestrogen does not produce a potentiation of the TAM-induced reduction of cell number, while the association of the drug with MPA seems more effective with respect to MPA alone, at least at certain concentrations. The negative interference observed between LND and TAM may be due to the LND-induced decrease of estrogen receptor levels.     

冬凌草甲素及联合柔红霉素对jurkat细胞增殖抑制作用的观察

目的：探讨冬凌草甲素单用及联合柔红霉素对急性T淋巴细胞白血病jurkat细胞增殖抑制的影响。方法：采用CCK一8法检测单用冬凌草甲素、柔红霉素对jurkat细胞的抑制作用,及两种药物联合应用对jurkat细胞的抑制作用;采用Giemsa法检测冬凌草甲素作用于jurkat细胞48h的凋亡形态学变化;采用流式细胞仪检测单用冬凌草甲素对jurkat细胞的凋亡率及两药物联合时的凋亡率。结果：单用不同浓度的冬凌草甲素（1、2、4、8和16μg／mL）对jurkat细胞具有增殖抑制作用,且呈时间-浓度依赖性,其中干预48h后的抑制率分别为（10．80±1．58）％、（32．32±7．83）％、（42．27±4．43）0、（55．07±1．65）％和（70．36±4．11）%;当联合小剂量柔红霉素（0．04μg／mL）时对jurkat细胞的抑制作用显著增加。Oiemsa染色后,随着药物浓度的增加,镜下可观察到胞体皱缩、染色质浓集和凋亡小体等形态学变化。不同浓度的冬凌草甲素干预jurkat细胞48h后的凋亡率分别为（7．74±0．96）％、（13．26士1．49）％、（17．42±1．24）％、（25．13±2．12）％和（29．07±0．59）％,呈浓度依赖性增加;单用柔红霉素（0．04μg／mL）干预jurkat细胞48h后的凋亡率为（18．19±1．33）％;当冬凌草甲素（4μg／mL）联合柔红霉素（0．04μg／mL）干预jurkat细胞时,其凋亡率为（51．06±2．25）％,与单用两种药物相比差异有统计学意义,P〈0．05。结论：冬凌草甲素能够抑制jurkat细胞增殖和诱导凋亡,联合柔红霉素对iurkat细胞的增殖抑制作用显著增强,具有协同抑制效应。     

In vitro activity of colistin as single agent and in combination with antifungals against filamentous fungi occurring in patients with cystic fibrosis

Because published reports indicate that the antibiotic colistin (COL) has antifungal properties, this study investigated the antifungal in vitro activity of COL as single agent and in combination with the antifungal compounds voriconazole (VRC), caspofungin (CAS) and amphotericin B (AMB) against Scedosporium/Pseudallescheria spp., Exophiala dermatitidis and Geosmithia argillacea. In total, susceptibility was determined for 77 Scedosporium/Pseudallescheria spp., 82 E. dermatitidis and 17 G. argillacea isolates. The minimal inhibitory concentrations (MICs) of COL and the antifungals as single compound and in combination were determined with MIC test strips. Drug interactions were detected by crossing the MIC test strips at a 90º angle. The fractional inhibitory concentration index was used to categorise the drugs’ interaction. The MIC₅₀ value of COL was 12 μg ml^?1 for S. prolificans, 16 μg ml^?1 for P. apiosperma, 16 μg ml^?1 for P. boydii, 12 μg ml^?1 for E. dermatiditis and 6 μg ml^?1 for G. argillacea. VRC was the most active drug in combination without any antagonism with the exception of few P. boydii isolates. COL as single agent and in most combinations with antifungals exhibits in vitro antifungal activity against filamentous ascomycetes occurring in cystic fibrosis patients and may offer a novel therapeutic option, especially for multidrug‐resistant S. prolificans.     

不同铂类为主联合化疗抗人肺癌细胞株的药效学研究*

探讨洛铂（ＬＢＰ）、顺铂（ＤＤＰ）和卡铂（Ｃａｂ）单药或联合紫杉醇（ＰＴＸ）、多西他赛（ＤＯＣ）和长春瑞滨（ＮＶＢ）抗人肺癌细胞株的作用和活性。方法　选用４种人非小细胞肺癌（ＮＳＣＬＣ）细胞株Ａ５４９、ＮＣＩ-Ｈ４６０、Ｃａｌｕ-３、ＮＣＩ Ｈ２３和小细胞肺癌（ＳＣＬＣ）细胞株ＮＣＩ-Ｈ５２６,用不同浓度的ＬＢＰ、ＤＤＰ和Ｃａｂ单独或与ＰＴＸ、ＤＯＣ和ＮＶＢ联合处理上述肺癌细胞株７２ｈ,应用磺酰罗丹明染色法评价细胞增殖,计算半数抑制浓度（ＩＣ_５０）。结果　ＬＢＰ、ＤＤＰ和Ｃａｂ单药均可抑制体外培养的肺癌细胞株生长,其中ＬＢＰ对４种ＮＳＣＬＣ细胞株的ＩＣ５０为１.７～２.４μｍｏｌ／Ｌ,ＤＤＰ为１.８～５.２μｍｏｌ／Ｌ,Ｃａｂ为２２.８～１００.８μｍｏｌ／Ｌ;ＬＢＰ、ＤＤＰ和Ｃａｂ对ＳＣＬＣ细胞株的ＩＣ５０分别为０.３μｍｏｌ／Ｌ、０.６μｍｏｌ／Ｌ和９.８μｍｏｌ／Ｌ。ＬＢＰ分别与ＰＴＸ、ＤＯＣ和ＮＶＢ联合抗ＮＳＣＬＣ细胞株有明显增效作用,其中与ＰＴＸ合用的协同作用最强。ＤＤＰ或Ｃａｂ分别与ＰＴＸ、ＤＯＣ和ＮＶＢ联合的抑制作用与细胞株的类型有关。结论　在上述３种铂类药物中,ＬＢＰ具有明确的抗ＮＳＣＬＣ细胞作用,与ＤＤＰ相似,而强于Ｃａｂ,其抗癌活性与细胞株的类型无关。ＬＢＰ分别联合ＰＴＸ、ＤＯＣ和ＮＶＢ均有协同作用,其中与ＰＴＸ的联合作用最强。     

Molecular characterization of Ambler class A to D β-lactamases,ISAba1, and integrons reveals multidrug-resistant Acinetobacter spp. isolates in northeastern China

The prevalence of various Ambler class A to D β-lactamases, ISAba1, and class 1 and 2 integrons as well as the clonal relatedness in 105 Acinetobacter spp. isolates found in northeastern China was investigated. All 105 Acinetobacter spp. isolates were determined to be multidrug resistant (MDR), and the resistance rates to carbapenem agents were approximately 50%. PER, IMP, AmpC, and OXA-23 were found to be dominant β-lactamases belonging to different classes, respectively. This is the first report of the coexistence of bla_PER, bla_IMP, bla_AmpC, and bla_OXA-23-like genes in Acinetobacter spp. isolates from northeastern China. ISAba1 was found upstream of the bla_OXA-23-like gene in 87.8% (36/41) strains and upstream of the bla_OXA-51-like gene in 26.5% (13/49) strains. ISAba3–like element was found upstream of the bla_OXA-58-like gene in one bla_OXA-58-like-positive strain. The presence of IntI1 was detected in 63.8% (67/105) of the isolates and the most prevalent gene cassettes were aacA4, aadA1, and catB8. The highly prevalent isolates belong to international clonal lineage (ICL)-II. These results indicate that the wide horizontal and clonal spread of MDR Acinetobacter spp. isolates harbouring multiple β-lactamase genes has become a serious problem in northeastern China.     

The effect of tirapazamine (SR-4233) alone or combined with chemotherapeutic agents on xenografted human tumours.

Recent data have shown that the in vitro and in vivo cytotoxicity of bioreductive drugs could be significantly increased when combined with chemotherapy drugs such as cisplatinum, depending on the timing of administration. The aim of this study was to define the toxicity (animal lethality) and the activity (growth delay assay, excision assay) of a bioreductive drug, tirapazamine, alone and combined with chemotherapy agents (5-FU, VP16, bleo, DTIC and c-DDP) on nude mice bearing xenografted human tumours: a rectal carcinoma (HRT18) and a melanoma (Na11+). Animal lethality was markedly increased when tirapazamine at the lethal dose 10% was combined with the other drugs. For the HRT18 tumour the combination of tirapazamine and bleomycin significantly increased the delay of regrowth compared with bleomycin alone (P = 0.04) and was more cytotoxic than tirapazamine alone (P = 0.04). For the Na11+ tumours the combination of tirapazamine with VP16 significantly increased tumour doubling time compared with the controls (P = 0.001) or VP16 alone. The combination of tirapazamine and VP16 was more cytotoxic than VP16 alone (P = 0.0001). When compared with c-DDP or tirapazamine alone, there was a significant decrease in plating efficiency when tirapazamine and c-DDP were given at the same time (P = 0.04), but not when tirapazamine was given 3 h before c-DDP. In conclusion, tirapazamine was shown to be cytotoxic against clonogenic human tumour cells. Its efficacy in vivo may depend on its combination with already active chemotherapy drugs on the tumour model used. The timing of administration may be less important than previously thought.     

嘌呤类似物单用或联合环磷酰胺治疗慢性淋巴细胞白血病安全性的Meta分析

目的系统评价嘌呤类似物联合环磷酰胺与单用嘌呤类似物治疗慢性淋巴细胞白血病（CLL）的安全性。方法检索PubMed数据库、ISI数据库、Embase数据库、Cochrane图书馆临床对照试验数据库、OVID数据库嘌呤类似物联合环磷酰胺、单用嘌呤类似物治疗CLL的随机对照试验,按Cochrance协作网推荐的方法进行质量评价、资料提取,采用Revman4．3．2软件进行Meta分析。结果共纳入4个随机对照试验,包括1358例患者。Meta分析结果显示,嘌呤类似物联合环磷酰胺与单用嘌呤类似物比较,严重白细胞减少（Ⅲ-Ⅳ级）发生率增加,差异有统计学意义[RR=1．47,95％置信区间（CI）1．10—1．98,P=0．011;严重贫血（Ⅲ—Ⅳ级）（RR=1．03,95％C10．74—1．43,P=0．87）、血小板减少（Ⅲ～Ⅳ级）（RR=1．28,95％CIO．99～1．65,P=0．06）发生率差异无统计学意义。结论与单用嘌呤类似物治疗CLL比较,嘌呤类似物联合环磷酰胺严重贫血、血小板减少发生率无明显增加,但严重白细胞减少发生率增加,临床使用时应引起重视。     

Effects of lonidamine alone or combined with hyperthermia in some experimental cell and tumour systems.

Lonidamine or 1-[(2, 4-dichlorophenyl) methyl]-1H-indazole-3-carboxylic acid, studied in a battery of in vitro and in vivo tests currently used for the screening of anti-tumour agents affecting cell division, has been shown to have a narrow spectrum of anti-tumour activity. The significance of this finding is discussed in the light of previous investigations suggesting that lonidamine affects mitochondrial function and not cell replication. Hyperthermia has been shown to sensitize tumour cells to lonidamine. This observation indicates that in combination with hyperthermia lonidamine has some potential for the treatment of cancer; moreover, it suggests that hyperthermia might reproduce a metabolic condition occurring in some stages of the disease. The blood levels corresponding to the anti-tumour action of lonidamine in animals are in the range of those detected in patients treated with the drug.     

Molecular mechanisms of resistance and toxicity associated with platinating agents

Platinating agents, including cisplatin, carboplatin, and oxaliplatin, have been used clinically for nearly 30years as part of the treatment of many types of cancers, including head and neck, testicular, ovarian, cervical, lung, colorectal and relapsed lymphoma. The cytotoxic lesion of platinating agents is thought to be the platinum intrastrand crosslink that forms on DNA, although treatment activates a number of signal transduction pathways. Treatment with these agents is characterized by resistance, both acquired and intrinsic. This resistance can be caused by a number of cellular adaptations, including reduced uptake, inactivation by glutathione and other anti-oxidants, and increased levels of DNA repair or DNA tolerance. Here we investigate the pathways that treatment with platinating agents activate, the mechanisms of resistance, potential candidate genes involved in the development of resistance, and associated clinical toxicities. Although the purpose of this review is to provide an overview of cisplatin, carboplatin, and oxaliplatin, we have focused primarily on preclinical data that has clinical relevance generated over the past five years.     

The effect of the hypoxic cell drug SR-4233 alone or combined with the ionizing radiations on two human tumor cell lines having different radiosensitivity.

The influence of the hypoxic cell drug, SR-4233, alone and/or combined with ionizing radiations on the survival of two human cell lines having very different intrinsic radiosensitivity was analysed. The killing effect of SR-4233 in hypoxia was similar for both cell lines. Twenty microM SR-4233 under hypoxic conditions had a killing effect equivalent to 6.6 Gy for the poorly sensitive cell line (HT29) and equivalent to 3.9 Gy for the highly sensitive cell line (SW48). The effect of SR-4233 and ionizing radiations on these two cell lines was tested in vitro: the cells were incubated for one hour in hypoxia with or without 20 microM SR-4233 and then irradiated in air. The HT29 cells that survived treatment with SR-4233 are more radiosensitive than untreated cells. However, their radiosensitivity is similar to that of cells that have been given a dose of 6.6 Gy. This suggests that SR-4233 acts additively, rather than as a radiosensitiser. As SR-4233 acts selectively in hypoxia, these results appear encouraging for the treatment of poorly-radiosensitive human tumors.     

沙利度胺治疗难治性复发性多发性骨髓瘤的临床研究

目的：观察沙利度胺(Thalidomide，国内商品名：反应停)单药或联合地塞米松治疗多发性骨髓瘤(Multiple Myeloma，MM)的疗效及副作用。方法：单药组：男性13例，女性2例，中位年龄58岁。其中2例为初发的MM；1例为原发性浆细胞白血病(PCL)；12例为难治性MM，其中3例为自体外周血干细胞移植(AutoPBSCT)术后复发。反应停治疗起始剂量为100mg／d，根据患者耐受情况，逐渐加量，最高达800mg／d。联合组：男性20例，女性7例，中位年龄56岁。其中初发1例；1例为原发性PCL；25例为难治MM，其中2例为自体外周血干细胞移植(AutoPBSCT)术后复发。反应停剂量为400mg／d左右加用地塞米松40mg／天，第1～4天，第9～12天，第17～20天，1个月为一个疗程。结果：单药组42．9％(6／14)的患者对治疗有效，其中3例为完全缓解(CR)或接近完全缓解(Near-CR)，1例有明显治疗反应(Major response)，1例为部分缓解(PR)。12例难治性MM中4例有效(33．3％)；联合组有效率为57．7％(15／26)，其中4例为CR或Near-CR，2例有明显治疗反应，9例为PR。25例难治性MM中11例有效(44．0％)。两组间在总有效率及难治病例的有效率方面均无显著差异。两组患者均出现不同程度的便秘、皮疹等副作用，但均可耐受。结论：沙利度胺单药或联合地塞米松对难治性复发性MM均有效。     

Comparative pharmacodynamic interaction analysis of triple combinations of caspofungin and voriconazole or ravuconazole with subinhibitory concentrations of amphotericin B against Aspergillus spp.

Triple combination therapy with an antifungal triazole, echinocandin and amphotericin B (AmB) is used in some centres to treat refractory aspergillosis. The objective of this study was to investigate the effect of subinhibitory concentrations of AmB on the double combinations of caspofungin (CAS) + voriconazole (VOR) or ravuconazole (RAV) against Aspergillus fumigatus, Aspergillus flavus and Aspergillus terreus. Isolates were studied in triplicate against CAS/VOR and CAS/RAV combinations by chequerboard broth microdilution. AmB was added to each double combination at concentrations of 0, 0.1 and 0.2 μg ml^?1. The fractional inhibitory concentration (FIC) index was calculated for the double and triple combinations. Comparative analysis was performed by repeated measures analysis followed by Dunnett’s post‐test. The double combinations of CAS/RAV and CAS/VOR were synergistic or additive in most conditions. Addition of AmB to the double combinations resulted in increased FIC indices for A. fumigatus and A. flavus. By contrast, AmB increased the synergism of the double combinations decreasing FIC indices for A. terreus (P < 0.05). RAV and VOR displayed similar synergistic activity with CAS. The addition of sub‐inhibitory amphotericin B concentrations reduced but did not eliminate the synergistic interaction between the echinocandin and triazole against A. fumigatus and A. flavus, while it increased the synergy against A. terreus.