Background The diagnosis of Clostridium difficile infection (CDI) increases concern that asymptomatic carriers of toxigenic C. difficile may be diagnosed with CDI. Methods A matched case control study was conducted in inpatients in a tertiary care center. The first 50 patients with diarrhea and a positive polymerase chain reaction (PCR) test beginning February 1, 2015, were identified as cases. Control patients were hospitalized patients receiving antibiotics, but with no diarrhea, housed in a room as close as possible to each case during the same admission time. A convenience sample of healthcare workers who cared for C. difficile infected patients was also tested. Results We found two positive PCR results for C. difficile in controls (4.1%). None of these healthcare workers were positive for C. difficile by PCR. There was no difference between groups with respect to overall antibiotic use before the requested PCR for Clostridium difficile (p = 0.359). The majority of cases had a high proportion of gastrointestinal disorders (71.4%) compared with control (8.2%), p < 0.001. Patients with neoplasia had a higher chance of being identified as cases (p = 0.041). Conclusions PCR should not be the only diagnostic tool but should be complementary to other methods and to the medical history.