ObjectiveTo investigate healthcareassociated infection(HAI) prevalence and antimicrobial use in a hospital. MethodsHAI prevalence rate, antimicrobial use and pathogen detection in all inpatients on August 21, 2013 were investigated with crosssectional survey method.  ResultsA total of 2 238 inpatients were investigated, 104 patients developed 126 times of HAI, HAI prevalence rate and case rate was 4.65% and 5.63% respectively; the top four sites for HAI were lower respiratory tract (28.57%), upper respiratory tract(18.25%), urinary tract(7.94%) and gastrointestinal tract(4.76%).Rate of specimens delivered for detection was 91.35%(95/104), the main specimen was sputum (26.32%),followed by blood (25.26%) and urine (10.53%). Antimicrobial usage rate was 24.58%, therapeutic, prophylactic plus therapeutic, and prophylactic use accounted for 36.55%, 45.09%, and 18.36% respectively; the usage rate of single, combination of 2, and 3 antimicrobial agents accounted for 75.82%,20.91% and 3.27% respectively. Risk factors of HAI were age (<15 or >60 years), all kinds of invasive procedures (tracheotomy, mechanical ventilation, urinary catheterization, arteriovenous  intubation, hemodialysis), and antitumor chemotherapy.  ConclusionSurvey on HIA prevalence helps to know the occurrence of HAI and usage of antimicrobial agents, as well as risk factors and highrisk departments of the occurrence of HAI, it also provides basis for subsequent targeted monitor on HAI.