Honors College Thesis
 

Studies on the evolution and expression of the Type II ferripyoverdine receptor gene fpvAIIb in Pseudomonas aeruginosa

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  • When iron levels are low, the opportunistic pathogen Pseudomonas aeruginosa secretes the high-affinity iron-binding siderophore pyoverdine. The cognate ferripyoverdine receptor, FpvA, has gained attention as the most divergent gene in the organism’s genome, showing substantial evidence of positive selection. S-type pyocins, bacteriocins with specificity to Pseudomonads, enter cells through FpvA and therefore may play a role in the evolution of these receptors. In order to study this interaction, an in vitro selection scheme was developed by testing the killing activity of pyocin S3 on P. aeruginosa ATCC 27853 under various conditions. A pyocin resistant mutant was isolated using the developed system. PCR analysis of the mutant revealed a whole-gene deletion of fpvA. This system provides a foundation for future fpvA evolution studies. In addition, these preliminary studies revealed important properties of pyocin S3. Killing activity of the pyocin was shown to be substantially more effective at 30°C than 37°C. Interestingly, pyocin S3 was also shown to kill ATCC 27853 even under iron-replete conditions, suggesting expression of fpvA is not regulated in concordance with iron availability in this strain. Real-time quantitative PCR (RT-PCR) was used to evaluate the expression of fpvAIIb in ATCC 27853 under different iron concentrations. mRNA concentrations were nearly equal under all iron conditions. Transcriptional regulation of pyoverdine receptor expression independently of iron concentration differs from the paradigm established with other Pseudomonas aeruginosa strains. Key Words: Pseudomonas aeruginosa, pyoverdine, pyocin, evolution, expression
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