Antimicrobial activity of spherical silver nanoparticles prepared using a biocompatible macromolecular capping agent: evidence for induction of a greatly prolonged bacterial lag phase.
Antimicrobial activity of spherical silver nanoparticles prepared using a biocompatible macromolecular capping agent: evidence for induction of a greatly prolonged bacterial lag phase.
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DOI：
10.1186/1477-3155-8-34
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发表时间：
2010-12-21
影响因子：
10.2
通讯作者：
Chen CY
中科院分区：
文献类型：
--
作者：
Irwin P;Martin J;Nguyen LH;He Y;Gehring A;Chen CY
我们用两种固相生物测定法对银基纳米粒子的抗菌性能进行了评估，发现10-20μL 0.3-3μM角蛋白稳定的纳米粒子(取决于起始细菌浓度=CI)完全抑制相当体积的金黄色葡萄球菌、鼠伤寒沙门氏菌或大肠杆菌O157：H7在固体表面上的生长。即使在37°C的固体培养上培养一周后，也没有观察到生长。在较低的NP浓度下(=[NP]S)，可以观察到可见的菌落，但它们最终停止生长。为了进一步研究这种生长抑制的生理学，我们在液体中重复这些实验，在37℃，在[NP]=0到10-6M的情况下，通过590 nm(OD)的光密度观察微生物的生长。为了提取各种生长参数，我们将所有OD[t]数据拟合到一个共同的S形函数，该函数提供开始和结束OD值的测量，一阶速率常数(K)，以及计算1/2-最大OD(Tm)的时间，这是CI，k的函数，以及微生物滞后时间(T)。使用96孔板阅读器进行这样的实验，我们发现生长总是发生在溶液中，但tm在7(对照组；CI=8×103CFU mL-1)和使用柠檬酸盐([NP]~3×10-7M)或角蛋白([NP]~10-6M)的20小时之间变化，并观察到{∂tm/∂[NP]}柠檬酸盐~5×107和{∂tm/∂[NP]}角蛋白~107小时·L摩尔-1。金黄色葡萄球菌的生长速率仅在k=1~1.2hr-1(1.1±0.075 hr-1)之间变化，对金黄色葡萄球菌的生长几乎没有影响。为了测试NPS正在改变细菌的初始浓度(CI)(即细胞死亡)的想法，我们进行了概率计算，假设tm中的扰动仅由于CI。我们发现，tm中如此大的扰动只能发生在任何增长概率都很小的CI中。这一结果表明，NP引起的tm的大部分变化是由于T显著增加(例如，从大约1小时增加到15-20小时)。对于固相分析，我们假设细菌最终变得不可培养，因为它们被抑制进行进一步的细胞分裂(T>；许多天)。我们认为，固体和液体系统之间的差异与NPS相对于细菌细胞膜的暴露或停留时间的明显差异有关，因为当选择小的NP抑制的菌落并在新鲜(即不存在NPS)上划线时，生长正常进行：例如，一个小的生长抑制的菌落在新鲜的固体介质上划线时会产生一大盘典型的金黄色葡萄球菌菌落。
We have evaluated the antimicrobial properties of Ag-based nanoparticles (Nps) using two solid phase bioassays and found that 10-20 μL of 0.3-3 μM keratin-stabilized Nps (depending on the starting bacterial concentration = CI) completely inhibited the growth of an equivalent volume of ca. 103 to 104 colony forming units per mL (CFU mL-1) Staphylococcus aureus, Salmonella Typhimurium, or Escherichia coli O157:H7 on solid surfaces. Even after one week at 37°C on solid media, no growth was observed. At lower Np concentrations (= [Np]s), visible colonies were observed but they eventually ceased growing. To further study the physiology of this growth inhibition, we repeated these experiments in liquid phase by observing microbial growth via optical density at 590 nm (OD) at 37°C in the presence of a [Np] = 0 to 10-6 M. To extract various growth parameters we fit all OD[t] data to a common sigmoidal function which provides measures of the beginning and final OD values, a first-order rate constant (k), as well as the time to calculated 1/2-maximal OD (tm) which is a function of CI, k, as well as the microbiological lag time (T). Performing such experiments using a 96-well microtitre plate reader, we found that growth always occurred in solution but tm varied between 7 (controls; CI = 8 × 103 CFU mL-1) and > 20 hrs using either the citrate-([Np] ~ 3 × 10-7 M) or keratin-based ([Np] ~ 10-6 M) Nps and observed that {∂tm/∂ [Np]}citrate ~ 5 × 107 and {∂tm/∂ [Np]}keratin ~ 107 hr·L mol-1. We also found that there was little effect of Nps on S. aureus growth rates which varied only between k = 1.0 and 1.2 hr-1 (1.1 ± 0.075 hr-1). To test the idea that the Nps were changing the initial concentration (CI) of bacteria (i.e., cell death), we performed probabilistic calculations assuming that the perturbations in tm were due to CI alone. We found that such large perturbations in tm could only come about at a CI where the probability of any growth at all was small. This result indicates that much of the Np-induced change in tm was due to a greatly increased T (e.g., from ca. 1 to 15-20 hrs). For the solid phase assays we hypothesize that the bacteria eventually became non-culturable since they were inhibited from undergoing further cell division (T > many days). We propose that the difference between the solid and liquid system relates to the obvious difference in the exposure, or residence, time of the Nps with respect to the bacterial cell membrane inasmuch as when small, Np-inhibited colonies were selected and streaked on fresh (i.e., no Nps present) media, growth proceeded normally: e.g., a small, growth-inhibited colony resulted in a plateful of typical S. aureus colonies when streaked on fresh, solid media.
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影响因子：
2.7
作者：
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