Document Type

Thesis

Degree Name

Master of Science (MSc)

Department

Biology

Program Name/Specialization

Integrative Biology

Faculty/School

Faculty of Science

First Advisor

Dr. Robin Slawson

Advisor Role

Supervisor

Abstract

Periodontitis is described as the inflammation of the periodontium and affects millions of people worldwide. This disease is caused by biofilm growth of certain bacteria on the tooth surface, adjacent to the gum tissue in the oral cavity. There are currently treatments against severe periodontitis, such as oral surgery and use of broad-spectrum antibiotics, but they fail to target the specific bacteria associated with periodontitis. Socransky et al., (1998), identified the preponderance for specific organisms in distinct areas of the subgingival pockets to associate with different bacteria forming consortia and complexes. The yellow complex, containing facultatively anaerobic, Gram positive Streptococcus oralis, Streptococcus mitis, Streptococcus gordonii, Streptococcus sanguinis and Streptococcus intermedius were the focus of this research. These bacteria form biofilms; enabling protection against the host immune system, sequestration of nutrients, and the formation of mixed communities. The streptococcal species that comprise the yellow complex contain adherence sites for other pathogenic bacteria, aiding in the progression of periodontal disease. This research focused on evaluating strategies and chemical efficacy to eliminate these organisms, which could lead to improved treatments against periodontitis. Yellow complex bacterial biofilms were analyzed both individually and as a multispecies biofilm. It was determined that under anaerobic conditions, with an absorbance of 2.3, the yellow complex form a slightly larger biofilm compared to aerobic conditions, with an absorbance of 2.1. Some members, S. intermedius and S. sanguinis, grew slower with a lower total cell biomass, an OD of approximately 1.0 under aerobic conditions, and 1.15 under anaerobic conditions, when compared to the other members with optical densities up to 1.4.Additionally, the multispecies biofilms were treated with two antimicrobial agents, hydrogen peroxide (0.01-10%) and sodium peroxide (5-55 mg/mL) using two techniques, co-inoculation and pre-formed biofilms. The minimum biofilm eradication concentration determined for co-inoculation varied between the aerobic and anaerobic environments. Key findings included that it was 7 mg/mL and 55 mg/mL of sodium fluoride that eradicated the yellow complex biofilms under aerobic conditions with pre-grown monoculture biofilms and pre-grown consortia biofilms, respectively, whereas, under anaerobic conditions, it was 55 mg/mL of NaF that eradicated the pre-grown monoculture biofilms, notably higher compared to exposure under aerobic conditions. Unfortunately, an MBEC value was not determined for pre-formed biofilms with the use of sodium fluoride. Unlike with sodium fluoride, the yellow complex biofilms had a higher resistance to hydrogen peroxide. The MBEC was determined to be 0.02% of H2O2 on pre-grown monocultures, but was not determined under any other conditions tested. Further, in an effort to more accurately identify the specific yellow complex members showing high resistance to the treatment compounds tested, primers were designed. It was found that the primers lacked uniqueness due to the similarities within the members genomes. Three of the primers tested, even though not unique, were specific to their target DNA. With this, standard curves, indicating number of gene copies per mL, were generated for some yellow complex members through qPCR. Ratios of each member within the biofilms were unable to be established for the yellow complex and, therefore, the population proportions were not determined. Ultimately, this research deepened our understanding of how the yellow complex members work together to form biofilms and, additionally, how the biofilms perform in the presence of an antimicrobial. This information can be used to improve the effectiveness of current treatments, such as the addition of antimicrobials and antibiotics to infected areas, which can aid in reducing the bacteria that cause periodontitis and its long-term effects.

Convocation Year

2020

Convocation Season

Fall

Available for download on Thursday, March 24, 2022

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