Alarming progressive increase in the prevalence of antimicrobial resistance in Escherichia coli has been documented worldwide. Previous studies have suggested that many E. coli clinical isolates are actually low-virulence opportunists whose success derives more from antibiotic resistance than from pathogenic capability. The co-existence of ESBL production and fluoroquinolone resistance was reported as a major therapeutic challenge for E. coli infections. Considering the sparse information regarding the genetic background of virulence and antibiotic resistance of local isolates, a collection of ciprofloxacin-resistant E. coli isolates from human extraintestinal specimens was analyzed using PCR, PCR-sequencing, and PFGE, in order to clarify some aspects regarding their mechanisms of antimicrobial resistance, phylogenetic origin, the content of virulence-encoding determinants, and clonal relatedness. The tested fluoroquinolone resistant E. coli (FQREC) isolates, which displayed genetic heterogeneity, carried double mutations in the QRDR of gyrA previously described, which could explain their high resistance to ciprofloxacin. More than half of them (69%) possessed group 1 blaCTX. like genes, and with one exception, all these isolates were ESBL producers. The FQREC isolates belonging to non B2 phylogenetic groups outnumbered the isolates derived from B2 group (60 versus 27 isolates), and their overall content of virulence-encoding genes (fim, pap, sfa/foc, afa, hly, cnf and aer) was reduced. Regardless of the phylogenetic origin, the most prevalent virulence-associated genes possessed by the FQREC isolates were aer and fim determinants, while none of these isolates carried hly and cnf genes. In the case of weakened patients, the E. coli isolates do not need a robust virulence repertoire in order to overcome the host defense systems. The co-resistance of many FQREC isolates to extended-spectrum cephalosporins may provide a substantial advantage to their survival and spreading within the hospital environment.