Signature-tagged mutagenesis (STM) is a genome-wide functional screening assay based upon an insertional mutation technique to identify mutants having a particular phenotype from a mixed population. The selection phenotype is either negative or positive for identification of mutations in genes which will impact the host as having a diminished or enhanced adaptation to a defined environment. STM has subsequently been used in different biological systems and adapted to use different mutagens, signature tags, and detection methods. Here, STM has been modified and used for genome-wide screening of Pseudomonas aeruginosa to identify genes and their products essential during the infection process or in enhancing virulence in vivo.