Title

Study of the Role of the msaABCR operon in Tricarboxylic Acid Cycle Activity and persister cell formation in Staphylococcus aureus

Date of Award

5-2021

Degree Type

Honors College Thesis

Academic Program

Biological Sciences BS

Department

Biological Sciences

First Advisor

Mohamed Elasri, Ph.D.

Advisor Department

Biological Sciences

Abstract

Staphylococcus aureus is an important human pathogen that causes wide arrays of infections ranging from minor skin infections to lethal systemic conditions such as infective endocarditis, osteomyelitis, sepsis and pneumonia. These systemic diseases are often difficult to treat due to the presence of persister cells. Persister cells are a phenotypic variant of the bacterial population that exhibit extreme and transient antibiotic tolerance accompanied by a transient halt in growth. Upon cessation of antibiotic treatment, however, persisters resume growth which results in recurrence of infections. This characteristic of persister cells therefore displays high clinical significance. In this study, we show the involvement of the msaABCR operon in antibiotic tolerance in S. aureus under the clinically relevant bactericidal antibiotic gentamicin. In previous study, the msaABCR operon was found to involve in persister cells formation such that the deletion of the msaABCR operon showed decreased persister formation in the methicillin resistant S. aureus (MRSA) strain USA300 LAC (Sahukhal et al. 2017). The RNA transcriptome of the msaABCR deletion mutant showed differential expression of genes that are involved in various metabolic pathways including carbohydrate, amino acid pathways (Sahukhal et al 2017). Considering the importance of metabolism in antibiotic tolerance, first we examined the expression of tricarboxylic acid (TCA) genes to measure the TCA cycle activity. The qRT-PCR results showed that the msaABCR deletion mutant indeed has increased expression of TCA genes and higher ATP content (Sahukhal et al., 2017). Furthermore, we evaluated the antibiotic tolerance in TCA cycle mutants in S. aureus. The results showed that the transposon mutants of TCA cycle genes exhibit higher antibiotic tolerance as compared to the wild type USA300 LAC. Considering the importance of energy content in the antibiotic tolerance, we also measured the ATP content and membrane potential. Deletion of TCA genes leads to decreased cellular ATP content as well as reduced membrane potential. We also observed increased pigment production and biofilm production in the TCA mutants. Altogether, these results suggest that the reduced metabolic activity contributes to the antibiotic tolerance as well as virulence in S. aureus.

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