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theses

ETD doctoral

Background: Antibiotic resistant Staphylococcus aureus (S. aureus) infections are a major medical concern due to loss of antibiotic sensitivity. Genome-wide analyses, including sequencing and gene expression, identified genes associated with antibiotic resistances, such as vraSR whose mutations are associated with vancomycin, daptomycin, and oxacillin resistances. Pathway enrichment analysis using Fisher’s Exact Test (FET) provides insight into pathway activity, though pathway roles in resistance are not fully elucidated. These studies applied a pathway-centric computational approach to examine antibiotic resistance (i.e., vraS-driven) and response (i.e., treatment inducible) changes in S. aureus.

Method: This is the first application of Gene Set Enrichment Analysis, which improves upon FET by removing gene selection requirements, to obtain pathway signatures (i.e., pathways ranked by activity change) from normalized enrichment scores reflecting 164 individual pathway activities in S. aureus. The pathway panels were obtained (most up- or down-regulated pathways separately), for vraS- and graSR-driven resistance signatures. A similar process was repeated to examine vancomycin susceptibility (i.e., difference in response between resistant and sensitive strains). Pathway activity in vraS-driven resistance panels was then examined in various antibiotic (vancomycin, oxacillin, or linezolid) susceptibilities to identify commonalities and differences in individual pathway activities. One novel pathway was selected and its association to antibiotic sensitivity was experimentally verified.

Results: This approach correlated pathway activity changes, like up-regulated histidine biosynthesis, with established genetic associations to antibiotic resistance. Further, pathway activity changes with known associations to vancomycin susceptibility such as down-regulated TCA cycle activity was also identified. Both examinations identified pathways with no prior association to antibiotic resistance. Inverse correlations between pathway activity changes and susceptibility to vancomycin and oxacillin/linezolid were seen (pathway activities up-regulated in vancomycin susceptibility were down-regulated in linezolid susceptibility) regardless of strain resistance level. Lysine biosynthesis was identified as a top candidate pathway for targeting to overcome resistance and verified by lysine or aspartate supplementation. Thus, lysine biosynthesis as a co-therapeutic target could restore antibiotic efficacy.

Conclusion: This pathway-centric approach identified pathway activity changes associated with antibiotic sensitivity which can be targeted to help reverse antibiotic resistance.

ETD_10953

doi:10.7282/t3-4mmy-q079

Ph.D.

Includes bibliographical references

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