Objective: To determine whether exposure to antimicrobial agents in the previous 90 days resulted in decreased bacterial susceptibility and increased hospital mortality in patients with severe sepsis or septic shock attributed to Gram-negative bacteremia. Design: A retrospective cohort study of hospitalized patients (January 2002 to December 2007). Setting: Barnes-Jewish Hospital, a 1200-bed urban teaching hospital. Patients: Seven hundred fifty-four consecutive patients with Gram-negative bacteremia complicated by severe sepsis or septic shock. Interventions: Data abstraction from computerized medical records. Measurements and Main Results: Escherichia coli (30.8%), Klebsiella pneumoniae (23.2%), and Pseudomonas aeruginosa (17.6%) were the most common isolates from blood cultures. Three hundred ten patients (41.1%) had recent antibiotic exposure. Cefepime was the most common agent with previous exposure (50.0%) followed by ciprofloxacin (32.6%) and imipenem or meropenem (28.7%). Patients with prior antibiotic exposure had significantly higher rates of resistance to cefepime (29.0% vs. 7.0%), piperacillin/tazobactam (31.9% vs. 11.5%), carbapenems (20.0% vs. 2.5%), ciprofloxacin (39.7% vs. 17.6%), and gentamicin (26.1% vs. 7.9%) (p < .001 for all comparisons). Patients with recent antibiotic exposure had greater inappropriate initial antimicrobial therapy (45.4% vs. 21.2%; p < .001) and hospital mortality (51.3% vs. 34.0%; p < .001) compared with patients without recent antibiotic exposure. Multivariate logistic regression analysis demonstrated that recent antibiotic exposure was independently associated with hospital mortality (adjusted odds ratio, 1.70; 95% confidence interval, 1.41–2.06; p = .005). Other variables independently associated with hospital mortality included use of vasopressors, infection resulting from P.aeruginosa, inappropriate initial antimicrobial therapy, increasing Acute Physiology and Chronic Health Evaluation II scores, and the number of acquired organ failures. Conclusions: Recent antibiotic exposure is associated with increased hospital mortality in Gram-negative bacteremia complicated by severe sepsis or septic shock. Clinicians caring for patients with severe sepsis or septic shock should consider recent antibiotic exposure when formulating empiric antimicrobial regimens for suspected Gram-negative bacterial infection.