Urinary tract infections (UTIs) are among the most frequent and significant bacterial infectious diseases, and are one of the greatest cost burdens on health systems worldwide. The recent emergence of antibitoic-resistant UTIs is a major concern, since untreatable infections can lead to kidney infection and potentially lethal sepsis. Uropathogenic E. coli (UPEC), which accounts for up to 90% of all UTIs, undergoes an intracellular infection cycle involving invasion and colonisation of cells of the bladder epithelium. Few genes essential to each phase of the UPEC infection cycle have been characterised. We aim to identify genes specifically required for the UPEC infection cycle in human bladder epithelial cells. First, a transposon-insertion mutant library of UPEC strain UTI-89 was prepared, to randomly inactivate UPEC genes on a genome-wide basis. The pooled library of mutants was then used to infect human bladder epithelial cells, replicating the phases of infection, in vitro. To identify mutants that were unable to proceed through the infection cycle, a next-generation DNA sequencing technique, known as transposon-directed insertion-site sequencing (TraDIS) is being used. Clones not present after infection would represent those genes essential for infection. This approach will be complemented with a bioinformatic search for E. coli genes that appear to be involved in the UPEC infection cycle and specific to UTIs. The combination of these two approaches will identify genes controlling UPEC infection of the human bladder.