Microbiology / Infectious Disease
Permanent URI for this collectionhttps://hdl.handle.net/20.500.12503/30816
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Browsing Microbiology / Infectious Disease by Author "Madunezim, Quentin"
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Item In vitro and in vivo development of rifampin resistance with Staphylococcus aureus clinical isolates(2022) Madunezim, QuentinPurpose: Rifampin is an older antibiotic used to treat several types of bacterial infections including tuberculosis. It is highly active against Staphylococcus aureus and effective against infections with this pathogen. Rifampin is a unique antibiotic that has the capacity to penetrate through thick bacterial biofilms and reach bacteria that may be harder to treat using other antibiotics. Rifampin's mechanism of action involves inhibiting bacterial RNA Polymerase by forming a stable drug-enzyme complex, however mutations in the rpoB gene that encodes the beta subunit of RNA polymerase can cause resistance in S. aureus. Resistant S.Aureus can then form biofilms on implanted medical devices such as prosthetic hips or indwelling catheters, making it even harder to treat and difficult on the lives of these individuals. The focus of our research was to determine if mutations in the rpoB gene of S. aureus induced in vitro leads to a decrease in pathogenicity of the bacteria, can be duplicated in an animal model during treatment and if rifampin can still be effective against these resistant mutants. Methods: The minimum inhibitory concentration (MIC) of rifampin was determined for several isolates of S. aureus using a broth microdilution method. These susceptible strains of S. aureus were then serially passaged on agar plates containing rifampin at multiples of the MIC. Colonies were selected from plates with the higher rifampin concentration, saved and then passaged again at increasing concentrations of rifampin. The change in rifampin MIC was confirmed for each isolate and passage by broth microdilution. The same serial passage was then conducted in broth. To confirm the purity, isolates were cultured on MSA plates to determine if contamination was present. Results: MICs for rifampin against the S. aureus isolates tested increased from 0.008 µg/mL for the parent strain to 16 µg/mL for selected strain after 3 passages on agar. Similar results were obtained following serial passaging in broth with rifampin MICs going from 0.002 µg/mL to 0.008 µg/mL. The observed increase in rifampin MIC for each isolate was found to be stable. Conclusion: Rifampin-resistant mutants were generated both in S. aureus 134-3 and 005-4 isolate strains following serial passages on agar plates and broth containing sub-inhibitory concentrations of rifampin. Rifampin MICs increased from parent to the resistant generated mutants (0.002 ug/mL to 16 ug/mL). Future studies will include looking at the same parent strains in an animal model of biofilm infection and demonstrating development of resistance to rifampin during therapy.