Date of Award

Spring 4-22-2024

Embargo Period

4-23-2024

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Regenerative Medicine and Cell Biology

College

College of Graduate Studies

First Advisor

Melinda Engevik

Second Advisor

Katherine Chetta

Third Advisor

Robin Muise-Helmericks

Fourth Advisor

Wei Jiang

Fifth Advisor

Ozlem Yilmaz

Abstract

Inflammatory Bowel Disease (IBD) is a chronic disorder characterized by excessive intestinal inflammation. Alterations in the gut microbiota have been consistently observed in individuals with IBD, but it is unclear which bacteria participate in driving inflammation. A comprehensive survey of publicly available RNA-sequencing datasets revealed that Acinetobacter species are elevated in the gastrointestinal tract of IBD patients. We found that A. calcoaceticus was particularly elevated in Crohn’s Disease patients; a subset of IBD. It is well documented that Acinetobacter species are resistant to several antibiotics, but there is very little information on the effects of Acinetobacter in the gut and hardly any data on A. calcoaceticus. Computational analysis of Acinetobacter genomes revealed that A. calcoaceticus harbored a wide repertoire of multi-drug efflux pump and beta-lactam resistance genes. We found that A. calcoaceticus strains were moderately to highly resistant to certain antibiotics within fluoroquinolones, aminoglycosides, tetracyclines, and other antibiotic classes. Modeling the conditions of the gut, we found that A. calcoaceticus strains tolerated varying levels of pH, osmolarity, ethanol and hydrogen peroxide. Biolog phenotypic microarrays further revealed that A. calcoaceticuscould use a range of nutrient sources found in the gut, including monosaccharides, disaccharides, glycosides, and amino acids. Moreover, A. calcoaceticus could grow anaerobically in stool-based bioreactors. Using inside-out intestinal organoids, we found that A. calcoaceticus stimulated pro-inflammatory cytokines; suggesting that A. calcoaceticus could initiate inflammation. In vivo, we found that A. calcoaceticus did not stimulate inflammation in the setting of a complex gut microbiota. However, disruption of the gut microbiota with antibiotics resulted in enhanced inflammation in A. calcoaceticus treated mice. To model IBD, we administered 2,4,6- trinitrobenzene sulfonic acid (TNBS) and we observed that TNBS treated mice receiving A. calcoaceticus lost more weight and had worse histological scores than mice treated with vehicle control and TNBS; indicating Acinetobacter worsens intestinal inflammation. Addition of antibiotics to the TNBS model further heightened the inflammation induced by A. calcoaceticus. Collectively, these data demonstrate that A. calcoaceticus is antibiotic resistant, well adapted to survive the gastrointestinal tract, and is capable of initiating inflammation in the setting of an altered gut microbiota.

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