Publication Date

6-15-2021

Document Type

Thesis

Degree Name

Master of Science in Biology (MS)

Committee Member

Combrink, Keith D.

Committee Member

Schmidl, Sebastian

Committee Member

Mendez, Monica

Abstract

Mycoplasma pneumoniae (MPN) causes community acquired pneumonia (CAP). The inability to identify M. pneumoniae infections through common clinical screenings has added to the global antibiotic resistance problem. MPN characteristically lacks a cell wall, which prevents proper diagnosis in an infected patient. This has led to the inappropriate prescriptions of antibiotics at insufficient dosages, allowing these microorganisms to thrive at sub-lethal concentrations. The small genome size of MPN, makes it a viable candidate for studying and developing tools to understand the genetic mechanisms behind the acquisition of antibiotic resistance by microbes. This study aimed to generate MPN mutants through prolonged exposure to 19 commercially available antibiotics. The objectives of this study were (1) to determine the minimum inhibitory concentration (MIC) of different antibiotic groups including macrolides, aminocyclitols, aminoglycosides, amphenicols, ansamycins, cephalosporins, fluoroquinolones, glycopeptides, ketolides, nitrofurans, and tetracyclines against M. pneumoniae (MPN) wild type strain M129-B7 and (2) to generate antibiotic resistant mutants through exposure of M. pneumoniae M129-B7 to constant sub-inhibitory (SIC) antibiotic concentrations across subsequent generations. MICs were determined qualitatively through color changes in the medium used for MIC assays. The averages of the MIC values indicate that the antibiotics: SPT, PUR, LVX, MXF, AZM, CLR, ERY, RXM, DOX, and TET are under the suggested breakpoint of ≤ 1 µg/mL, which supports the efficacy of these drugs against unaltered versions of MPN M129-B7. Alternatively, GEN, KAN, CHL, RFB, CFZ, CRO, CIP, CLI, and NIT had MIC averages higher than 1 µg/mL. These MIC values were used to generate four separate sets of passages under decreasing sub-inhibitory concentrations (SIC). The antibiotics: SPT, GEN, KAN, PUR, CHL, CIP, LVX, MXF, CLI, DOX, and TET were tested on MPN M129-B7 and successfully generated five passages under antibiotic pressure for each. This study shows that antibiotic pressure from SPT, GEN, KAN, CIP, and MXF antibiotics can generate mutations in MPN M129-B7 within five passages. This indicates that we were able to further develop techniques to use on M. pneumoniae as a model system to lead to the exploration of the genetic mechanisms involved in antibiotic resistance development in mycoplasmas and possibly other microbes.

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