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The importance of growth kinetic analysis in determining bacterial susceptibility against antibiotics and silver nanoparticles.

Theophel K, Schacht VJ, Schlüter M, Schnell S, Stingu CS, Schaumann R, Bunge M - Front Microbiol (2014)

Bottom Line: Despite the delayed onset of growth, treatment with fosfomycin, daptomycin, fusidic acid, cefoxitin, or gentamicin resulted in higher maximum growth rates and/or higher final optical density values compared with antibiotic-free controls, indicating that growth stimulation and hormetic effects may occur with extended exposure to sublethal antibiotic concentrations.Whereas neither maximum growth rate nor final cell density correlated with antibiotic concentration, the lag phase duration for some antibiotics was a more meaningful indicator of dose-dependent growth inhibition.Our results also reveal that non-temporal growth profiles are only of limited value for cultivation-based antimicrobial silver nanoparticle susceptibility testing.

View Article: PubMed Central - PubMed

Affiliation: Institute of Applied Microbiology, Research Center for BioSystems, Land Use, and Nutrition, Justus Liebig University of Giessen Giessen, Germany.

ABSTRACT
Routine antibiotics susceptibility testing still relies on standardized cultivation-based analyses, including measurement of inhibition zones in conventional agar diffusion tests and endpoint turbidity-based measurements. Here, we demonstrate that common off-line monitoring and endpoint determination after 18-24 h could be insufficient for reliable growth-dependent evaluation of antibiotic susceptibility. Different minimal inhibitory concentrations were obtained in 20- and 48 h microdilution plate tests using an Enterococcus faecium clinical isolate (strain UKI-MB07) as a model organism. Hence, we used an on-line kinetic assay for simultaneous cultivation and time-resolved growth analysis in a 96-well format instead of off-line susceptibility testing. Growth of the Enterococcus test organism was delayed up to 30 h in the presence of 0.25 μg mL(-1) of vancomycin and 8 μg mL(-1) of fosfomycin, after which pronounced growth was observed. Despite the delayed onset of growth, treatment with fosfomycin, daptomycin, fusidic acid, cefoxitin, or gentamicin resulted in higher maximum growth rates and/or higher final optical density values compared with antibiotic-free controls, indicating that growth stimulation and hormetic effects may occur with extended exposure to sublethal antibiotic concentrations. Whereas neither maximum growth rate nor final cell density correlated with antibiotic concentration, the lag phase duration for some antibiotics was a more meaningful indicator of dose-dependent growth inhibition. Our results also reveal that non-temporal growth profiles are only of limited value for cultivation-based antimicrobial silver nanoparticle susceptibility testing. The exposure to Ag(0) nanoparticles led to plasma membrane damage in a concentration-dependent manner and induced oxidative stress in Enterococcus faecium UKI-MB07, as shown by intracellular ROS accumulation.

No MeSH data available.


Related in: MedlinePlus

Inhibition of growth of the E. faecium isolate by simultaneous treatment with antibiotics and silver nanoparticles. Results for only ciprofloxacin and moxifloxacin are shown, but inhibitory effects were observed with all antibiotics tested. For visual clarity, SD (n = 3, error bars) are only presented for selected time points.
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Figure 2: Inhibition of growth of the E. faecium isolate by simultaneous treatment with antibiotics and silver nanoparticles. Results for only ciprofloxacin and moxifloxacin are shown, but inhibitory effects were observed with all antibiotics tested. For visual clarity, SD (n = 3, error bars) are only presented for selected time points.

Mentions: The monochromator-based instrument used in this study enables adjustment of the optical measurement settings in the presence of dispersed nanoparticles. Metal nanoparticles often exhibit strong background signals, which can affect absorbance measurements due to nanoparticle-specific (e.g., metal speciation and particle size distribution) and concentration-dependent properties. To examine putative synergism between antibiotics and metal nanoparticles, we compared the growth characteristics of the Enterococcus isolate in the presence of antibiotics or nanoparticles alone with its growth in the presence of both antibiotics and Ag(0) nanoparticles. Although growth was only partially inhibited by antibiotics alone at the tested concentrations (see Figure 1), the organism did not grow in the presence of both antibiotics and nanoparticles. The inhibitory effects of combined antibiotic/Ag(0) nanoparticle treatments involving ciprofloxacin and moxifloxacin are shown in Figures 2A,B.


The importance of growth kinetic analysis in determining bacterial susceptibility against antibiotics and silver nanoparticles.

Theophel K, Schacht VJ, Schlüter M, Schnell S, Stingu CS, Schaumann R, Bunge M - Front Microbiol (2014)

Inhibition of growth of the E. faecium isolate by simultaneous treatment with antibiotics and silver nanoparticles. Results for only ciprofloxacin and moxifloxacin are shown, but inhibitory effects were observed with all antibiotics tested. For visual clarity, SD (n = 3, error bars) are only presented for selected time points.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4226228&req=5

Figure 2: Inhibition of growth of the E. faecium isolate by simultaneous treatment with antibiotics and silver nanoparticles. Results for only ciprofloxacin and moxifloxacin are shown, but inhibitory effects were observed with all antibiotics tested. For visual clarity, SD (n = 3, error bars) are only presented for selected time points.
Mentions: The monochromator-based instrument used in this study enables adjustment of the optical measurement settings in the presence of dispersed nanoparticles. Metal nanoparticles often exhibit strong background signals, which can affect absorbance measurements due to nanoparticle-specific (e.g., metal speciation and particle size distribution) and concentration-dependent properties. To examine putative synergism between antibiotics and metal nanoparticles, we compared the growth characteristics of the Enterococcus isolate in the presence of antibiotics or nanoparticles alone with its growth in the presence of both antibiotics and Ag(0) nanoparticles. Although growth was only partially inhibited by antibiotics alone at the tested concentrations (see Figure 1), the organism did not grow in the presence of both antibiotics and nanoparticles. The inhibitory effects of combined antibiotic/Ag(0) nanoparticle treatments involving ciprofloxacin and moxifloxacin are shown in Figures 2A,B.

Bottom Line: Despite the delayed onset of growth, treatment with fosfomycin, daptomycin, fusidic acid, cefoxitin, or gentamicin resulted in higher maximum growth rates and/or higher final optical density values compared with antibiotic-free controls, indicating that growth stimulation and hormetic effects may occur with extended exposure to sublethal antibiotic concentrations.Whereas neither maximum growth rate nor final cell density correlated with antibiotic concentration, the lag phase duration for some antibiotics was a more meaningful indicator of dose-dependent growth inhibition.Our results also reveal that non-temporal growth profiles are only of limited value for cultivation-based antimicrobial silver nanoparticle susceptibility testing.

View Article: PubMed Central - PubMed

Affiliation: Institute of Applied Microbiology, Research Center for BioSystems, Land Use, and Nutrition, Justus Liebig University of Giessen Giessen, Germany.

ABSTRACT
Routine antibiotics susceptibility testing still relies on standardized cultivation-based analyses, including measurement of inhibition zones in conventional agar diffusion tests and endpoint turbidity-based measurements. Here, we demonstrate that common off-line monitoring and endpoint determination after 18-24 h could be insufficient for reliable growth-dependent evaluation of antibiotic susceptibility. Different minimal inhibitory concentrations were obtained in 20- and 48 h microdilution plate tests using an Enterococcus faecium clinical isolate (strain UKI-MB07) as a model organism. Hence, we used an on-line kinetic assay for simultaneous cultivation and time-resolved growth analysis in a 96-well format instead of off-line susceptibility testing. Growth of the Enterococcus test organism was delayed up to 30 h in the presence of 0.25 μg mL(-1) of vancomycin and 8 μg mL(-1) of fosfomycin, after which pronounced growth was observed. Despite the delayed onset of growth, treatment with fosfomycin, daptomycin, fusidic acid, cefoxitin, or gentamicin resulted in higher maximum growth rates and/or higher final optical density values compared with antibiotic-free controls, indicating that growth stimulation and hormetic effects may occur with extended exposure to sublethal antibiotic concentrations. Whereas neither maximum growth rate nor final cell density correlated with antibiotic concentration, the lag phase duration for some antibiotics was a more meaningful indicator of dose-dependent growth inhibition. Our results also reveal that non-temporal growth profiles are only of limited value for cultivation-based antimicrobial silver nanoparticle susceptibility testing. The exposure to Ag(0) nanoparticles led to plasma membrane damage in a concentration-dependent manner and induced oxidative stress in Enterococcus faecium UKI-MB07, as shown by intracellular ROS accumulation.

No MeSH data available.


Related in: MedlinePlus