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Date of Award

Spring 2004

Document Type

Restricted Thesis: Campus only access

Degree Name

Bachelor of Science



First Advisor

Dr. Laura Runyen-Janecky


The intracellular pathogen Shigella flexneri is a highly infectious bacterium that causes shigellosis, or dysentery, in humans. During invasion of human cells, S. flexneri induces the expression of numerous genes to respond to the host cell environment. One such set of genes, the pst operon, is predicted to encode a high affinity phosphate acquisition system (Pst) that is activated under conditions of reduced phosphate. Previous work has shown that a mutation in the pst operon causes S. flexneri to form smaller plaques (zones of killing) in human cell monolayers than the normal strain. Studies have also shown that the Pi-regulated gene phoA is expressed at elevated levels in the pst mutant as compared to the normal strain. This suggests that the inability of the S. flexneri pst mutant to form wild-type plaques in Henle cell monolayers may be due to abnormal activation of Pi-regulated genes such as phoA. To test this hypothesis, a mutation in phoB, the gene encoding the regulator activated by the Pst system, was constructed in the S. flexneri pst mutant to eliminate expression of Pi-regulated genes such as phoA. In this pst/phoB mutant, the phoB mutation suppressed the small plaque phenotype seen with the pst mutant. Additionally, a specific mutation (R220Q) was constructed in the pstA gene of the pst operon that was predicted to eliminate Pst-mediated phosphate transport but have normal Pi-regulated gene expression, based on the phenotype of an Escherichia coli strain harboring the same mutation. When this pstAR220Q mutation was added back into a S. flexneri pst mutant, the resultant pstAR220Q strain exhibited normal plaque formation, as well as normal regulation of phoA expression. It was also found, however, that the initial pst mutation did not affect S. flexneri's resistance to acidic environments, which could have had important implication for S. flexneri's growth and survival in vivo. These findings provide further support for the model of abnormal expression of Pi-regulated genes negatively affecting plaque formation, and indicate that the regulatory function of the Pst system in S. flexneri is of greater significance to its plaque formation than is the system's high affinity phosphate transport function.