Uropathogenic <i>Escherichia coli</i> (UPEC) is a major bacterial pathogen that causes urinary tract infections (UTIs). The mouse is an available UTI model for studying the pathogenicity; however, <i>Caenorhabditis elegans</i> represents as an alternative surrogate host with the capacity for high-throughput analysis. Then, we established a simple assay for a UPEC infection model with <i>C. elegans</i> for large-scale screening. A total of 133 clinically isolated <i>E. coli</i> strains, which included UTI-associated and fecal isolates, were applied to demonstrate the simple pathogenicity assay. From the screening, several virulence factors (VFs) involved with iron acquisition (<i>chuA</i>, <i>fyuA,</i> and <i>
irp2</i>) were significantly associated with high pathogenicity. We then evaluated whether the VFs in UPEC were involved in the pathogenicity. Mutants of <i>E. coli</i> UTI89 with defective iron acquisition systems were applied to a solid killing assay with <i>C. elegans</i>. As a result, the survival rate of <i>C. elegans</i> fed with the mutants significantly increased compared to when fed with the parent strain. The results demonstrated, the simple assay with <i>C. elegans</i> was useful as a UPEC infectious model. To our knowledge, this is the first report of the involvement of iron acquisition in the pathogenicity of UPEC in a <i>C</i>. <i>elegans</i> model.