Role of aggregative adherence fimbriae from enteroaggregative Escherichia coli isolates in biofilm and colonization

Enteroaggregative Escherichia coli (EAEC) are a diverse group of bacteria that cause diarrhea worldwide. EAEC significantly affect travelers to endemic regions, including military personnel, and children in developing countries where EAEC infection is associated with childhood failure-to-thrive. EAEC creates thick biofilms on the intestinal mucosa, a process that is thought to contribute to the development of both diarrhea and childhood failure-to-thrive. Typical EAEC strains encode and produce just one aggregative adherence fimbriae (AAF) out of the five different AAF types. The AAF are required for aggregative adherence to epithelial cells in vitro, but the degree of importance of each of the AAF types in both biofilm formation and pathogenesis is unknown. In this study, we investigated the role of the fimbriae in EAEC biofilms by deleting the major fimbrial subunit gene for the AAF from each of the five AAF categories and observing the impact on biofilm staining from recent EAEC clinical isolates. We found that biofilm was significantly reduced in all strains when the AAF gene was deleted, and that the defect could be overcome by complementation. In this work we also describe a modified murine EAEC model appropriate for colonization studies. In an antibiotic-treated mouse colonization model, some AAF mutant strains were attenuated for colonization, including AAF/II, AAF/IV, and AAF/V isolates. We did not observe complementation of the attenuated colonization phenotype in the mouse model. However, since we found a colonization defect for several EAEC mutant strains of different AAF types, a link between the fimbriae and colonization in the mice is supported. Taken together, our results show that the AAF are required for biofilm formation, and that some AAF contribute to colonization in a mouse model.