Abstract Background Escherichia coli bloodstream infections (BSIs) account for high mortality rates (5%–30%). Determinants of death are unclear, especially since the emergence of ESBL producers. Objectives To determine the relative weight of host characteristics, bacterial virulence and antibiotic resistance in the outcome of patients suffering from E. coli BSI. Methods All consecutive patients suffering from E. coli BSI in seven teaching hospitals around Paris were prospectively included for 10 months. E. coli isolates were sequenced using Illumina NextSeq technology to determine the phylogroup, ST/ST complex (STc), virulence and antimicrobial resistance gene content. Risk factors associated with death at discharge or Day 28 were determined. Results Overall, 545 patients (mean ± SD age 68.5 ± 16.5 years; 52.5% male) were included. Mean Charlson comorbidity index (CCI) was 5.6 (± 3.1); 19.6% and 12.8% presented with sepsis and septic shock, respectively. Portals of entry were mainly urinary (51.9%), digestive (41.9%) and pulmonary (3.5%); 98/545 isolates (18%) were third-generation cephalosporin resistant (3GC-R), including 86 ESBL producers. In-hospital death (or at Day 28) was 52/545 (9.5%). Factors independently associated with death were a pulmonary portal of entry [adjusted OR (aOR) 6.54, 95% CI 2.23–19.2, P = 0.0006], the iha_17 virulence gene (aOR 4.41, 95% CI 1.23–15.74, P = 0.022), the STc88 (aOR 3.62, 95% CI 1.30–10.09, P = 0.014), healthcare-associated infections (aOR 1.98, 95% CI 1.04–3.76, P = 0.036) and high CCI (aOR 1.14, 95% CI 1.04–1.26, P = 0.006), but not ESBL/3GC-R. Conclusions Host factors, portal of entry and bacterial characteristics remain major determinants associated with mortality in E. coli BSIs. Despite a high prevalence of ESBL producers, antibiotic resistance did not impact mortality. (ClinicalTrials.gov identifier: NCT02890901.)