Chiou C C, Mavrogiorgos N, Tillem E, Hector R, Walsh T J
Immunocompromised Host Section, Pediatric Oncology Branch, National Cancer Institute, Bethesda, Maryland 20892, USA.
Antimicrob Agents Chemother. 2001 Dec;45(12):3310-21. doi: 10.1128/AAC.45.12.3310-3321.2001.
We investigated the potential synergy between two cell wall-active agents, the echinocandin FK463 (FK) and the chitin synthase inhibitor nikkomycin Z (NZ), against 16 isolates of filamentous fungi. Susceptibility testing was performed with a broth macrodilution procedure by NCCLS methods. The median minimal effective concentration (MEC) of FK against all Aspergillus species was 0.25 microg/ml (range, 0.05 to 0.5 microg/ml). For Fusarium solani and Rhizopus oryzae, MECs of FK were >512 microg/ml. The median MEC of NZ against Aspergillus fumigatus was 32 microg/ml (range, 8 to 64 microg/ml), and that against R. oryzae was 0.5 microg/ml (range, 0.06 to 2 microg/ml); however, for the other Aspergillus species, as well as F. solani, MECs were >512 microg/ml. A checkerboard inhibitory assay demonstrated synergy against A. fumigatus (median fractional inhibitory concentration index = 0.312 [range, 0.15 to 0.475]). The effect was additive to indifferent against R. oryzae and indifferent against other Aspergillus spp. and F. solani. We further investigated the pharmacodynamics of hyphal damage by MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay and examined the time-sequenced changes in hyphal ultrastructure. Significant synergistic hyphal damage was demonstrated with the combination of NZ (2 to 32 microg/ml) and FK (0.03 to 0.5 microg/ml) over a wide range of concentrations (P < 0.001). The synergistic effect was most pronounced after 12 h of incubation and was sustained through 24 h. Time-sequenced light and electron microscopic studies demonstrated that structural alterations of hyphae were profound, with marked transformation of hyphae to blastospore-like structures, in the presence of FK plus NZ, while fungi treated with a single drug showed partial recovery at 24 h. The methods used in this study may be applicable to elucidating the activity and interaction of other cell wall-active agents. In summary, these two cell wall-targeted antifungal agents, FK and NZ, showed marked time-dependent in vitro synergistic activity against A. fumigatus.
我们研究了两种细胞壁活性药物棘白菌素FK463（FK）和几丁质合成酶抑制剂多氧霉素Z（NZ）对16株丝状真菌的潜在协同作用。采用美国国家临床实验室标准化委员会（NCCLS）方法通过肉汤稀释法进行药敏试验。FK对所有曲霉菌种的中位最小有效浓度（MEC）为0.25微克/毫升（范围为0.05至0.5微克/毫升）。对于茄病镰刀菌和米根霉，FK的MEC>512微克/毫升。NZ对烟曲霉的中位MEC为32微克/毫升（范围为8至64微克/毫升），对米根霉的中位MEC为0.5微克/毫升（范围为0.06至2微克/毫升）；然而，对于其他曲霉菌种以及茄病镰刀菌，MEC>512微克/毫升。棋盘抑制试验证明对烟曲霉有协同作用（中位分数抑制浓度指数=0.312[范围为0.15至0.475]）。对米根霉的作用为相加至无作用，对其他曲霉菌种和茄病镰刀菌为无作用。我们通过MTT[3-(4,5-二甲基噻唑-2-基)-2,5-二苯基四氮唑溴盐]试验进一步研究了菌丝损伤的药效学，并检查了菌丝超微结构的时间序列变化。在很宽的浓度范围内（P<0.001），NZ（2至32微克/毫升）和FK（0.03至0.5微克/毫升）联合使用显示出显著的协同菌丝损伤。协同作用在孵育12小时后最为明显，并持续至24小时。时间序列的光学和电子显微镜研究表明，在FK加NZ存在的情况下，菌丝的结构改变很显著，菌丝明显转变为芽生孢子样结构，而用单一药物处理的真菌在24小时时显示部分恢复。本研究中使用的方法可能适用于阐明其他细胞壁活性药物的活性和相互作用。总之，这两种针对细胞壁的抗真菌药物FK和NZ在体外对烟曲霉显示出显著的时间依赖性协同活性。
