Background: The Clostridium difficile rate in symptomatic patients represents both those with C difficile infection (CDI) and those with colonization. To predict the extent of CDI overdiagnosis, we compared the asymptomatic colonization rate to the symptomatic positivity rate in hospitalized patients using nucleic acid testing. Methods: Between July 2014 and April 2015, formed stool samples were collected from asymptomatic patients after admission to 3 hospital wards at the Stanford Hospital. Stool samples from symptomatic patients with suspected CDI in the same wards were collected for testing per provider order. The GeneXpert C. difficile tcdB polymerase chain reaction (PCR) assay (Cepheid, Sunnyvale, CA, USA) was performed on all stool samples and PCR cycle threshold was used as a measure of genomic equivalents. Chart review was performed to obtain clinical history and medication exposure. Results: We found an asymptomatic C. difficile carriage rate of 11.8% (43/365) (95% confidence interval [CI], 8.515.1%) and a positivity rate in symptomatic patients of 15.4% (54/351) (95% CI, 11.6-19.2%; P = 0.19). The median PCR cycle thresholds was not significantly different between asymptomatic carriers and symptomatic positives (29.5 versus 27.3; P = 0.07). Among asymptomatic patients, 11.6% (5/43) of carriers and 8.4% (27/322; P = 0.56) of noncarriers subsequently became symptomatic CDI suspects within the same hospitalization. Single and multivariate analysis did not identify any demographic or clinical factors as being significantly associated with C difficile carriage. Conclusions: Asymptomatic C difficile carriage rate was similar to symptomatic positivity rate. This suggests the majority of PCR-positive results in symptomatic patients are likely due to C. difficile colonization. Disease specific biomarkers are needed to accurately diagnose patients with C difficile disease. (C) 2017 Elsevier Inc. All rights reserved.