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Gold Nanoparticle-Targeted Laser Therapy for Enhancement of Antibiotic Efficacy against MRSA Biofilms



One novel technology that addresses this treatment need involves the use of pulsed laser energy to disperse the biofilm matrix and, resultantly, remove the barrier to penetration of antibiotics to the bacterial cells residing within the biofilm. When combined with gold nanoparticles (GNPs) coated with anti-bacterial antibodies, the laser energy can be harnessed and localized to the area of the infection. This strategy takes advantage of the excellent energy-absorbing capacity of GNPs, the amenability of GNPs to functionalize with a wide range of targeting moieties, and the ability of lasers at specific wavelengths to deliver light energy to the GNPs.
Irradiation of GNPs bound to the surface of the bacteria with a nanosecond pulsed laser causes extremely rapid heating (photothermal effect) and, in some cases, vapor bubble formation followed by cooling, contraction, and generation of acoustic waves (optoacoustic effect) around the particles.11 These phenomena lead to thermal and mechanical damage to the extracellular matrix and bacteria and, ultimately, dispersal of the biofilm. Elicitation of these effects around the GNPs occurs on a nanosecond time scale, providing very rapid destruction of the biofilm. Because the underlying damage mechanisms are physical (thermal and mechanical) rather than chemical (antibiotic-based), development of resistance is less likely, and this therapy can be combined with traditional antibiotics for potentially additive or synergistic effects.
In addition, it has been hypothesized that use of very short laser pulses and targeted GNPs localizes the effects to the biofilm and, in this way, may minimize collateral host tissue damage.11 Other advantages of this therapy are that it involves the use of laser parameters currently employed in clinical practice, and that GNPs have demonstrated good biocompatibility during testing in clinical trials.12
Successful demonstration of the efficacy of this experimental technology against methicillin-resistant Staphylococcus aureus (MRSA) infections was recently presented by researchers from the Naval Medical Research Unit San Antonio at the ASM Microbe 2016 Conference held June 16-20, 2016, in Boston. GNP-targeted laser therapy was tested as a potential strategy to destroy and disperse biofilms using an in-vitro MRSA infection model. In this work, GNPs were first functionalized with antibodies specific for S. aureus peptidoglycan, a component of the bacterial cell wall.
MRSA biofilms were grown for 24 hours then pretreated with various concentrations of antibody-coated GNPs for 2 hours. Unattached GNPs were washed away, and the biofilms were then irradiated using a nanosecond pulsed laser with a wavelength of 532 nm. Following laser irradiation, a subset of biofilms was exposed to gentamycin for 24 hours. Samples were then analyzed by laser scanning confocal microscopy and colony-forming unit assays to determine the extent of biofilm dispersal and bacterial cell viability after treatment.
Data revealed that treatment of biofilms with GNP-assisted laser therapy alone or increasing concentrations of gentamycin alone for 24 hours resulted in a maximum reduction in bacterial cell viability of only 1-log compared with control samples. However, exposure of biofilms to combination therapy of GNP-targeted pulsed laser therapy followed by 24-hour gentamycin treatment led to a 4-log reduction in viable bacteria. Thus, combining the GNP-assisted laser therapy and antibiotic treatment led to a synergistic enhancement of effect against the MRSA biofilms. Notably, this 4-log reduction in the viability of the biofilms was similar to that observed for gentamycin alone against planktonic MRSA cultures, suggesting that the GNP-assisted laser therapy dispersed the biofilm to allow access of the antibiotic to act against the bacteria. Confocal microscopy images (Figure 1) clearly indicate a zone of damage and confirm that the GNP-targeted laser therapy dispersed the MRSA biofilms. Additionally, biofilms treated with pulsed laser irradiation alone or nontargeted (uncoated) GNPs plus laser exposure remained unaffected, with no significant reduction in biofilm biomass or cell viability compared with the control samples. This indicates the antibody targeting strategy does, in fact, lead to increased binding of GNPs and localization of pulsed laser energy to the biofilm.

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