Autotransporters (AT) are widespread in Gram-negative bacteria, and many of them are involved in virulence. An open reading frame (APECO1_O1CoBM96) encoding a novel AT was located in the pathogenicity island of avian pathogenicEscherichia coli (APEC) O1's virulence plasmid, pAPEC-O1-ColBM. This 3.5-kb APEC autotransporter gene (aatA) is predicted to encode a 123.7-kDa protein with a 25-amino-acid signal peptide, an 857-amino-acid passenger domain, and a 284-amino-acid β domain. The three-dimensional structure of AatA was also predicted by the threading method using the I-TASSER online server and then was refined using four-body contact potentials. Molecular analysis of AatA revealed that it is translocated to the cell surface, where it elicits antibody production in infected chickens. Gene prevalence analysis indicated that aatA is strongly associated with E. coli from avian sources but not with E. coli isolated from human hosts. Also, AatA was shown to enhance adhesion of APEC to chicken embryo fibroblast cells and to contribute to APEC virulence.