objective. To compare incidence of hospital-onset Clostridium difficile infection (CDI) measured by the use of International Classification of Diseases, Ninth Revision, Clinical Modification (ICD-9-CM) discharge diagnosis codes with rates measured by the use of electronically available C. difficile toxin assay results. Methods. Cases of hospital-onset CDI were identified at 5 US hospitals during the period from July 2000 through June 2006 with the use of 2 surveillance definitions: positive toxin assay results (gold standard) and secondary ICD-9-CM discharge diagnosis codes for CDI. The x 2 test was used to compare incidence, linear regression models were used to analyze trends, and the test of equality was used to compare slopes. Results. Of 8,670 cases of hospital-onset CDI, 38% were identified by the use of both toxin assay results and the ICD-9-CM code, 16% by the use of toxin assay results alone, and 45% by the use of the ICD-9-CM code alone. Nearly half (47%) of cases of CDI identified by the use of a secondary diagnosis code alone were community-onset CDI according to the results of the toxin assay. The rate of hospitalonset CDI found by use of ICD-9-CM codes was significantly higher than the rate found by use of toxin assay results overall (P <.001), as well as individually at 3 of the 5 hospitals (P <.001 for all). The agreement between toxin assay results and the presence of a secondary ICD-9-CM diagnosis code for CDI was moderate, with an overall k value of 0.509 and hospital-specific k values of 0.489-0.570. Overall, the annual increase in CDI incidence was significantly greater for rates determined by the use of ICD-9-CM codes than for rates determined by the use of toxin assay results (Pp.006). Conclusions. Although the ICD-9-CM code for CDI seems to be adequate for measuring the overall CDI burden, use of the ICD-9-CM discharge diagnosis code for CDI, without present-on-admission code assignment, is not an acceptable surrogate for surveillance for hospital-onset CDI.