The efflux inhibitor phenylalanine-arginine beta-naphthylamide (PAβN) permeabilizes the outer membrane of gram-negative bacteria.
The efflux inhibitor phenylalanine-arginine beta-naphthylamide (PAβN) permeabilizes the outer membrane of gram-negative bacteria.
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DOI：
10.1371/journal.pone.0060666
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发表时间：
2013
期刊：
影响因子：
3.7
通讯作者：
Burrows LL
中科院分区：
文献类型：
--
作者：
Lamers RP;Cavallari JF;Burrows LL
抗菌药物的主动外排是细菌病原体产生多重耐药的主要机制。外排泵抑制剂（EPI）与泵底物的联合使用正在探索中，以克服外排介导的多药耐药。苯丙氨酸-精氨酸β-萘酰胺（PAβN）是一种研究充分的EPI，通常与氟喹诺酮类抗生素联合使用，但很少有研究评估其与β-内酰胺类抗生素联合使用的效用。本研究的最初目标是评估β-内酰胺类药物与PAβN联合治疗条件致病菌铜绿假单胞菌的疗效。PAβN降低了几种β-内酰胺类抗生素对铜绿假单胞菌的最小抑菌浓度（MIC）;然而，敏感性变化并不完全是由于外排抑制。PAβN处理后，使β-内酰胺抗生素失活的染色体编码AmpC β-内酰胺酶的细胞内水平显著降低，上清液中AmpC水平相应增加，可能是由于外膜的透化作用。PAβN处理导致8-苯胺基-1-萘磺酸（一种荧光疏水探针）的摄取显著增加，并使铜绿假单胞菌对通常不能穿过外膜的大体积抗生素（如万古霉素）以及去污剂样胆汁盐敏感。在生长培养基中补充镁以稳定外膜，可增加PAβN存在下的MIC，并恢复对万古霉素的耐药性。因此，PAβN以浓度依赖性方式渗透细菌膜，其水平低于联合研究中通常使用的水平，当使用PAβN作为外排研究的对照时，应考虑这种额外的作用模式。
Active efflux of antimicrobial agents is a primary mechanism by which bacterial pathogens can become multidrug resistant. The combined use of efflux pump inhibitors (EPIs) with pump substrates is under exploration to overcome efflux-mediated multidrug resistance. Phenylalanine-arginine β-naphthylamide (PAβN) is a well-studied EPI that is routinely combined with fluoroquinolone antibiotics, but few studies have assessed its utility in combination with β-lactam antibiotics. The initial goal of this study was to assess the efficacy of β-lactams in combination with PAβN against the opportunistic pathogen, Pseudomonas aeruginosa. PAβN reduced the minimal inhibitory concentrations (MICs) of several β-lactam antibiotics against P. aeruginosa; however, the susceptibility changes were not due entirely to efflux inhibition. Upon PAβN treatment, intracellular levels of the chromosomally-encoded AmpC β-lactamase that inactivates β-lactam antibiotics were significantly reduced and AmpC levels in supernatants correspondingly increased, potentially due to permeabilization of the outer membrane. PAβN treatment caused a significant increase in uptake of 8-anilino-1-naphthylenesulfonic acid, a fluorescent hydrophobic probe, and sensitized P. aeruginosa to bulky antibiotics (e.g. vancomycin) that are normally incapable of crossing the outer membrane, as well as to detergent-like bile salts. Supplementation of growth media with magnesium to stabilize the outer membrane increased MICs in the presence of PAβN and restored resistance to vancomycin. Thus, PAβN permeabilizes bacterial membranes in a concentration-dependent manner at levels below those typically used in combination studies, and this additional mode of action should be considered when using PAβN as a control for efflux studies.
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