Defense Date

7-15-2020

Document Type

Thesis

Degree Type

Master of Science

Degree Name

Biological Sciences

First Advisor

Robert Smith, Ph.D.

Second Advisor

Julie T. Garcia, Ph. D.

Third Advisor

Omar T. Eldakar, Ph. D.

Abstract

Infections due to bacteria were once easily treated using antibiotics. However, the effective shelf life of antibiotics is diminishing due to the rise and spread of antibiotic resistant bacteria. Accordingly, novel approaches to treating infections are required. Pseudomonas aeruginosa is a multi-drug resistant, biofilm forming opportunistic pathogen. A determinant of P. aeruginosa’s pathogenicity is pyoverdine, a siderophore that is used to sequester iron from the environment. After being produced by a bacterium, pyoverdine is secreted into the environment where it complexes with iron. The pyoverdine-iron complex is then returned to the bacteria through diffusion, where it drives the synthesis of additional pyoverdine. Accordingly, the relative positions of bacteria and pyoverdine are important to facilitate access to this pyoverdine-iron complex. Pyoverdine is critically important in the infection process as its removal or attenuation results in decreased infection severity. Interestingly, previous studies have suggested that periodically disrupting spatial structure of a bacterial population, through physical means, can disrupt the ability of bacteria to access diffusible molecules required for growth. Based on this previous work, we hypothesized that disrupting biofilms composed of P. aeruginosa biofilms using physical disturbance would prevent accessibility to pyoverdine, which would ultimately reduce its synthesis. To test this hypothesis, we periodically disturbed the spatial structure of a biofilm using a microplate reader. We found that a disturbance frequency of 6 shakes/hr, we were able to significantly reduce endpoint pyoverdine amount compared to the control frequencies of 0 shakes/hr and 30 shakes/hr. This observation was consistent when the density of biofilms was increased using medium with increasing concentrations of glucose. Interestingly, we observed periodic disturbance at 6 shake/hr could also disturb the ability of 20 different P. aeruginosa strains isolated from the clinic to produce pyoverdine; in some cases, the amount of pyoverdine increased, decreased, or stayed the same as a result of periodic disturbance. Overall, this suggests that the ability to perturb pyoverdine production through periodic disturbance is not limited to a single parameter space (one strain, one growth rate, one biofilm density), and instead may be observable in strains and environments that lead to diverse biofilm densities. As pyoverdine is required for the infection process, it is possible that our research might lead to novel strategies to reduce, or prevent, infections due to P. aeruginosa.

Available for download on Thursday, August 05, 2021

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