The spread of carbapenem-resistant members of the Enterobacteriaceae family (CRE) harboring carbapenemases is an emerging public health threat. As KPC-producing Klebsiella species are endemic in our tertiary care hospital, we aimed to evaluate a PCR-based surveillance test for identification of rectal carriage of KPC-producing CRE. We conducted a surveillance study between May and December 2007. Rectal swabs were collected from patients known to harbor CRE and from contacts of newly discovered patients harboring CRE. Specimens were evaluated by culture and by PCR analysis for blaKPC and were defined as positive if CRE was cultured and blaKPC was identified. Discrepant results between the culture and PCR analysis were resolved by subculturing, repeating the PCR, and performing a hydrolysis assay. Positive CRE cultures prior or subsequent to the time of sampling for the study were also taken into consideration. Sensitivity, specificity, and time to result were calculated. A total of 755 swabs were included. Concordant results were documented for 735 specimens; 51 were positive as determined by both PCR and culture. Discrepancies existed for 20 swabs; 9 were blaKPC negative and CRE culture positive, and 11 were blaKPC positive and CRE culture negative. After repeat testing, a total of 64 samples were classified as blaKPC-positive CRE. The sensitivity and specificity of the PCR analysis were 92.2% and 99.6%, respectively, and those of the culture were 87.5% and 99.4%, respectively. Over the last 3 months of the study, the sensitivity of the PCR improved to 96.3%, versus 77.8% for culture. Time to result was 30 h for the PCR and 60 h (negative) and 75 h (positive) for the CRE culture. blaKPC PCR-based testing is a useful method for the surveillance of KPC-producing CRE. Its main advantage over culturing is a shorter time to result, and it may prove to be more sensitive.