Date of Award

Spring 5-2021

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

School

Biological, Environmental, and Earth Sciences

Committee Chair

Dr. Mohamed O. Elasri

Committee Chair School

Biological, Environmental, and Earth Sciences

Committee Member 2

Dr. Glenmore Shearer

Committee Member 2 School

Biological, Environmental, and Earth Sciences

Committee Member 3

Dr. Fengwei Bai

Committee Member 3 School

Biological, Environmental, and Earth Sciences

Committee Member 4

Dr. Janet R. Donaldson

Committee Member 4 School

Biological, Environmental, and Earth Sciences

Committee Member 5

Dr. Shahid Karim

Committee Member 5 School

Biological, Environmental, and Earth Sciences

Abstract

Staphylococcus aureus is a major human pathogen that causes lethal systemic conditions such as infective endocarditis, osteomyelitis, sepsis and pneumonia. Chronic in nature, these diseases are often refractory to the antibiotic treatment. Such recalcitrance is mainly caused due to the presence of persister cells, which are a bacterial subpopulation that exhibits extreme, yet transient antibiotic tolerance accompanied by a transient halt in the growth. However, upon cessation of antibiotic treatment, resumption in growth of persister cells causes recurrence of infections and treatment failure, displaying tremendous clinical significance. In this study, we show the involvement of the msaABCR operon in persister cells formation in S. aureus under clinically relevant bactericidal antibiotics. Considering the importance of metabolism in antibiotic tolerance, we demonstrate causal link between cellular energy and antibiotic tolerance in S. aureus. We observe that the inactivation of the msaABCR operon in USA300 LAC led to increased tricarboxylic acid (TCA) cycle activity, cellular ATP and NADH content, and membrane potential. The increased ATP in the msaABCR mutant during late exponential-phase led to decreased persister formation against different antibiotic stress, while the increased membrane potential in the stationary-phase led to defective persister formation against aminoglycoside stress. Furthermore, we show msaABCR (MsaB) as a transcriptional repressor of catabolite control protein E (ccpE), an activator of TCA cycle, and nicotinamide dehydrogenase (ndh2), the gene of electron transport chain (ETC) in S. aureus.

Deletion of the msaABCR operon resulted in the downregulated expression of several genes involved in resistance against oxidative stress. Notably, those included carotenoid biosynthetic genes (crtM/N), OsmC/Ohr family protein (ohr) and catalase (katA) genes. Here, we report the transcriptional regulation of the crtOPQMN operon and the ohr gene by the msaABCR operon. In addition, we also show MsaB as the transcriptional activator of sigB (transcriptional regulator of the crtOPQMN operon) and transcriptional repressor of sarZ (repressor of ohr). These results suggest that the msaABCR operon regulates an oxidative-stress defense mechanism, which is required to facilitate persistent and recurrent staphylococcal infections. In conclusion, we demonstrate the importance of the msaABCR operon in S. aureus survival in different stressful environment.

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