Inpatients in the intensive care unit (ICU) are at high risk for healthcare-associated infections (HAIs). In the current study, a bundle of interventions and measures for preventing and controlling HAIs were developed and implemented in the ICU by trained personnel, and the impact of the bundle was evaluated. The incidence of HAIs, the adjusted daily incidence of HAIs and the incidence of three types of catheter-related infections before and after the bundle implementation were compared. The execution rate of the bundle for preventing and controlling ventilator-associated pneumonia (VAP) was increased from 82.06% in 2012 to 96.88% in 2013. The execution rate was increased from 83.03% in 2012 to 91.33% in 2013 for central line-associated bloodstream infection (CLABSI), from 87.00% to 94.40% for catheter-associated urinary tract infection (CAUTI), and from 82.05% to 98.55% for multidrug-resistant organisms (MDROs), respectively. In total, 136 cases (10.37%) in 2012 and 113 cases (7.72%) in 2013 involved HAIs, respectively. Patients suffered from infection of the lower respiratory tract, the most common site of HAIs, in 134 cases (79.29%) in 2012 and 107 cases (74.30%) in 2013 respectively. The incidence of VAP was 32.72‰ and 24.60‰, the number of strains of pathogens isolated was 198 and 173, and the number of MDROs detected in the ICU was 91 and 74 in 2012 and 2013, respectively. The percentage of MDROs among the pathogens causing HAIs was decreased in each quarter of 2013 as compared with the corresponding percentage in 2012. In 2013, the execution rate of the bundle for preventing and controlling HAIs was increased, whereas the incidence of HAIs and VAP decreased as compared with that in 2012.