Extended-spectrum β-lactamases (ESBL) are widespread enzymes in animals, and the risk of transmission of ESBL genes to humans has become a major issue. In Tunisia, recent data showed a high prevalence of ESBL-producing Escherichia coli isolates in healthy animals, mostly in chickens. In this study, we report the first data on ESBL in diseased Tunisian animals (chickens and cattle), highlighting a major difference in ESBL prevalence in the infectious versus noninfectious E. coli flora. Interestingly, the only ESBL producer was an ST10 E. coli from a cattle, and not from chicken. Moreover, this E. coli isolate harbored the bla(CTX-M-15) gene on an F2:A-:B- plasmid, a combination frequently found in humans. This plasmid was also highly similar to a bla(CTX-M-15) F2:A-:B- plasmid recently reported in cattle in France. Altogether, this study is also the first report of the bla(CTX-M-15) gene in food animals in Tunisia, and, to our best knowledge, the first report of an ESBL producer in cattle in Africa. Since this plasmid was recognized in cattle in France and worldwide in humans, the question of its origin in Tunisian cattle is open. The detection of ESBL producers in milk in Tunisia may also constitute a risk of ESBL transmission from animals to humans through food consumption.