Abstract 1483 Investigating the Metabolic Effects of Sulfur Limitation in Pseudomonas aeruginosa PAO1

Pseudomonas aeruginosa PAO1 is a ubiquitous pathogen that is notoriously prevalent in healthcare-associated infections (HAIs). As a multi-drug resistant organism, it is difficult to treat and can counteract host immune clearance during infections through the expression of multiple virulence factors. Preliminary studies suggest that sulfur limitation affects the ability of P. aeruginosa PAO1 to express specific virulence factors. Sulfur is an essential element that can be utilized by P. aeruginosa for metabolic activity, with inorganic sulfur being the preferred source. However, inorganic sulfur is limiting in the environment, so there are alternative mechanisms that P. aeruginosa PAO1 can use to acquire and assimilate organosulfur sources into usable forms. The primary global transcriptional regulator responsible for alternative sulfur assimilation in P. aeruginosa PAO1 is the CysB protein. CysB is transcribed from the cys operon of P. aeruginosa PAO1 during sulfur limiting conditions and upregulates enzymes and proteins essential for the uptake and assimilation of organosulfur compounds. Since an adequate sulfur supply is required for optimal bacterial virulence, it is hypothesized that CysB may play an indirect regulatory role in promoting bacterial pathogenicity. Therefore, our group evaluated how different virulence properties of P. aeruginosa PAO1 were affected by the deletion of CysB (ΔCysB) relative to the wild-type organism. P. aeruginosa PAO1 and the ΔCysB variant showed similar growth curves and colorimetric evidence of biofilm formation in sulfur-free media supplemented with L-cysteine. In comparison studies with P. aeruginosa PAO1, the ΔCysB variant portrayed similar biofilm formation, but showed decreased growth when supplemented with sulfate. Interestingly, limited biofilm formation and growth was observed for both the P. aeruginosa PAO1 and the ΔCysB variant when grown under sulfur-limiting conditions. In addition to biofilm and growth studies, proteomic mass spectrometry was performed on the planktonic and sessile state cells to identify how the different growth conditions directly altered protein expression. The findings from these studies suggests that sulfur starvation diminishes the expression of virulence factors in P. aeruginosa PAO1 and deletion of the CysB regulator exacerbates decreased growth. Additionally, the cumulative data indicates the shift from the planktonic to sessile stage is dependent on alternative pathways for sulfur assimilation.