Date of Award

12-2009

Document Type

Thesis

Degree Name

Master of Science (MS)

Legacy Department

Environmental Toxicology

Advisor

Klaine, Stephen J

Committee Member

McNealy , Tamara L

Committee Member

Bain , Lisa

Abstract

There exists widespread concern of pathogenic bacteria colonizing and establishing biofilms in water systems providing a direct exposure route to the public. Legionella are widely distributed in human-made and natural environments, and colonization lends the potential to infect human hosts that may lead to the development of Legionnaire's Disease. Silver has been used in cooling towers and water filtration systems for removal of planktonic bacteria and biofilms; however permanent removal of L. pneumophila biofilms with metal ions is impossible when established biofilms are in hard to access places such as dead end piping or inside a protozoan host. Bulk metals showed limited success in the past at the permanent removal of Legionella , but nanometals, unlike their bulk elemental or molecular form, behave based on their size, shape and surface chemistries. Some nanometals may provide, perhaps, an interesting avenue for novel anti-microbial agents. This research investigated interactions between citrate-capped gold nanoparticles (AuNPs) and L. pneumophila biofilms. Planktonic virulence, growth kinetics and toxicity were assessed, while biofilms were evaluated for structure, density, and composition. Transmission electron microscopy (TEM) micrographs indicate that AuNPs associate with cellular surfaces, the internal cellular compartment and integrate into the extrapolymeric matrix. Previous research has shown that Au and Ag NPs induce an antibacterial response in some planktonic bacteria. In contrast, there were no significant differences of planktonic viability of L. pneumophila treated with 18 nm AuNPs. However, planktonic assays with 4 nm AuNPs at a concentration of 100 µg/L indicated a statistically significant decrease in biomass, measured as the optic density (OD) at 600 nm after 30 hrs of growth. Pigment production, a known virulence factor, measured after 48 hrs of growth at OD400 for the 4nm AuNPs treatments was also significantly less than all other treatments. Evaluation of the biofilm using Syto 11 dye and confocal microscopy indicate an alteration of morphology seen on glass slides. In treated biofilms, L. pneumophila formed long chain-like assemblages in contrast to a thick and densely packed biofilm as seen in controls indicating a possible stress response to the presence of AuNPs. No change in morphology was seen between controls and polystyrene NP treatments. AuNPs, although not effective in completely inhibiting L. pneumophila growth and pigment production, exhibit a negative mechanical response under biofilm forming conditions in response to 18 and 4 nm AuNPs. The results from this research suggest the potential uses of NPs in destabilization of biofilms. Further research will attempt to delineate whether the effects from surface chemistries, sizes, or shapes of the AuNPs are merely mechanical or antimicrobial with L. pneumophila .

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Biology Commons

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