Background: The rise of infections caused by multidrug-resistant Gram negative bacilli (MDR-GNB), added to paucity of newer therapy, have led to increase polymyxin B use, despite adverse renal toxicity profile.Aim: To determine the incidence and risk factors associated to acute kidney injury (AKI) and polymyxin B use, in patients with infections caused by MDR-GNB.Methods: A retrospective cohort, with a nested case-control study of adults who received polymyxin B for more than 48 hours at a tertiary university hospital in Colombia (2011-2015) was performed. AKI was definedby AKIN criteria. Results: Of 139 patients included in our study, 102 were male with median age of 49 years (IQR:28-64). Sixty-one patients (44%) developed AKI. Independent risk factors for development of AKI included: total polymyxin B daily dose (OR = 2.19, 95% CI, 1.04-4.64); length of stay at ICU (OR = 1.03, 95% CI, 1.00-1.06); nosocomial infection (OR = 6.43, 95% CI, 2.12, -19.47); and vasopressor use (OR = 5.38, 95% CI, 2.40-12.07). Mortality was higher among AKI-patients (58.6%) compared with non-AKI patients (25.6%) (p = 0.001). Conclusion: In this study, the rate of AKI associated to polymyxin B use was greater than reported in studies from last decade, and associated with increased mortality. AKI associated to polymyxin B use is likely multifactorial and aggravated by the critically ill state of patients sufferingnosocomial infections caused by mdr-gnb.