Summary Quaternary ammonium compounds (QACs) are cationic surfactants used as preservatives and environmental disinfectants. Limited data are available regarding the effect of QACs in the clinical setting. We performed a prospective cohort study in 153 patients with Escherichia coli bacteraemia from February to September 2008 at University Hospital in Rennes. The minimum inhibitory concentrations (MICs) of antibiotics and QACs alkyldimethylbenzylammonium chloride (ADBAC) and didecyldimethylammonium chloride (DDAC) were determined by the agar dilution method. The capacity of biofilm production was assayed using the Crystal Violet method, and mutation frequencies by measuring the capacity of strains to generate resistance to rifampicin. Logistic regression analysis showed that one of the significant factors related to low MICs for ADBAC (≤16 mg/L) and DDAC (≤8 mg/L), was cotrimoxazole susceptibility (odds ratio: 3.72; 95% confidence interval: 1.22–11.24; P = 0.02 and OR: 3.61; 95% CI: 1.56–7.56; P < 0.01, respectively). Antibiotic susceptibility to cotrimoxazole was strongly associated with susceptibility to amoxicillin and nalidixic acid ( P < 0.01). Community-acquired or healthcare-associated bacteraemia, severity of bacteraemia, and patient outcome were independent of the MICs of ADBAC and DDAC. Our findings demonstrate an epidemiological relationship between higher MIC values of QACs in clinical E. coli isolates and antibiotic resistance.