Date on Master's Thesis/Doctoral Dissertation

12-2022

Document Type

Doctoral Dissertation

Degree Name

Ph. D.

Department

Biology

Degree Program

Biology, PhD

Committee Chair

Yoder-Himes, Deborah

Committee Co-Chair (if applicable)

Menze, Michael

Committee Member

Menze, Michael

Committee Member

Perlin, Michael

Committee Member

Potempa, Jan

Committee Member

Goldberg, Joanna

Author's Keywords

Staphylococcus aureus; Burkholderia cenocepacia; microbial ecology; biofilms; antimicrobial; antibiotic

Abstract

Antimicrobial resistance is a phenomenon of increasing concern as antimicrobial overuse and misuse eliminate current therapeutic options, ushering society into a post-antimicrobial era. Antibiotic discovery and synthesis efforts are urgently needed to counter the increasing burden of antimicrobial resistance. Staphylococcus aureus is a causative agent of a variety of clinical manifestations including bacteremia, endocarditis, soft tissue infection, osteomyelitis, and device-related infections. S. aureus infection presents additional treatment challenges due to its capacity for biofilm formation, which is a mode of growth that confers protection from antibiotics and physical elimination, and the emergence of antibiotic resistant strains, including methicillin-resistant S. aureus and vancomycin-resistant S. aureus. Infection with antibiotic-resistant strains occurs within both nosocomial and community settings, broadening the potential impact of this organism. Bacteria within the genus Burkholderia hold vast potential as sources of antimicrobial agents. Our analysis of patient culture data, provided by the Cystic Fibrosis Foundation, suggests a negative relationship between members of the Burkholderia cepacia complex and Staphylococcus aureus. An in vitro screen for activity against S. aureus indicated several clinical strains of Burkholderia cenocepacia confer potent anti-Staphylococcus activity. This dissertation characterizes the deleterious effect of the presence of B. cenocepacia J2315 and H111, two clinical isolates from cystic fibrosis patients, against S. aureus. Co-culture biofilm-associated survival of both methicillin-sensitive and methicillin-resistant strains was, overall, decreased with both B. cenocepacia J2315 and H111. I further established the breadth of antibiotic activity of these two strains in co-culture with multiple Staphylococcus and other Gram-positive species, including Enterococcus, Bacillus and Listeria. While both B. cenocepacia strains demonstrated detrimental effects against survival of co-inoculated Staphylococcus species, the extent of inhibition of other Gram-positive species differed. Antagonistic activity against the Enterococcus and Bacillus strains assessed in co-culture with B. cenocepacia H111 was profound, with reduction of many co-cultured strains to below the limit of detection. Co-culture survival of the same Gram-positive species with B. cenocepacia J2315 indicated no significant reduction versus cognate mono-culture. Inhibition of S. aureus by both B. cenocepacia strains occurs via a secreted compound, as evidenced by reduction in survival of S. aureus when exposed to B. cenocepacia sterile biofilm supernatants. The inhibitory substance, at least for B. cenocepacia J2315 is secreted in larger quantities in response to the presence of S. aureus. Enzymatic treatment of the supernatants suggests that a protein and an RNA, or a nucleoprotein, are involved in the B. cenocepacia J2315-mediated antagonism of S. aureus, but that inhibition by B. cenocepacia H111 involves a different mechanism. The inhibitory effect is largely dependent upon culture medium, and B. cenocepacia J2315 is more sensitive to differences in nutrient composition of the growth medium than B. cenocepacia H111. Further, this decrease in viable S. aureus is not simply due to B. cenocepacia causing a release of S. aureus from biofilms but is due to killing of S. aureus. Collectively, these data confirm the biotechnological potential of two B. cenocepacia strains and serve to optimize conditions for observation and analysis of this phenomenon.

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