Chlorhexidine oral care is widely used in critically and non-critically ill hospitalized patients to maintain oral health. We investigated the effect of chlorhexidine oral care on mortality in a general hospitalized population. In this single-center, retrospective, hospital-wide, observational cohort study we included adult hospitalized patients (2012-2014). Mortality associated with chlorhexidine oral care was assessed by logistic regression analysis. A threshold cumulative dose of 300 mg served as a dichotomic proxy for chlorhexidine exposure. We adjusted for demographics, diagnostic category, and risk of mortality expressed in four categories (minor, moderate, major, and extreme). The study cohort included 82,274 patients of which 11,133 (14%) received chlorhexidine oral care. Low-level exposure to chlorhexidine oral care (≤ 300 mg) was associated with increased risk of death [odds ratio (OR) 2.61; 95% confidence interval (CI) 2.32-2.92]. This association was stronger among patients with a lower risk of death: OR 5.50 (95% CI 4.51-6.71) with minor/moderate risk, OR 2.33 (95% CI 1.96-2.78) with a major risk, and a not significant OR 1.13 (95% CI 0.90-1.41) with an extreme risk of mortality. Similar observations were made for high-level exposure (> 300 mg). No harmful effect was observed in ventilated and non-ventilated ICU patients. Increased risk of death was observed in patients who did not receive mechanical ventilation and were not admitted to ICUs. The adjusted number of patients needed to be exposed to result in one additional fatality case was 47.1 (95% CI 45.2-49.1). These data argue against the indiscriminate widespread use of chlorhexidine oral care in hospitalized patients, in the absence of proven benefit in specific populations.