Avian pathogenic Escherichia coli (APEC) is the etiological agent of avian colibacillosis, a leading cause of economic loss to the poultry industry. APEC has enhanced survivability due to biofilm formation. To better characterize APEC, this work evaluated E. coli isolated from a turkey cellulitis outbreak. Using PCR, the O serogroups of the isolates were identified, and it was found that O143, a potential emerging APEC serogroup, may be a leading cause of the outbreak. The next study aimed to better understand virulence factors that contribute to APEC biofilm formation. Signature tagged mutagenesis was used to ultimately identify nine APEC- and biofilm-associated genes. Deletion mutants were created to evaluate the role of these genes in APEC biofilm formation. In conclusion, novel genes not previously known to be APEC- or biofilm-associated were identified. This work builds the foundation for future studies to characterize these genes and better understand APEC biofilm.