We evaluated the clinical performance of Check-Direct CPE for carbapenemase detection directly from 301 perirectal swabs (258 patients) in a nonoutbreak setting. Culture of a PCR-confirmed, carbapenemase-containing organism, or history of colonization with such organism within the previous 2 weeks, was used as the reference standard. Check-Direct CPE demonstrated a sensitivity value, specificity value, positive predictive value (PPV), and negative predictive value (NPV) of 100% (all bla KPC ), 88%, 21%, and 100%, respectively. False positives accounted for 79% (n ؍ 34) of samples for which a cycle threshold (C T ) value was reached. Simulated studies to evaluate specimen pooling as an approach to minimize costs showed no difference in C T values for pooled groups of three or five that each contained a single specimen spiked with ϳ1,500 CFU bla KPC Klebsiella pneumoniae; however, the detection rate dropped to 60% at a seeded concentration of ϳ150 CFU. When data were pooled, C T values for bla KPC were higher for heavy-feces-containing than for light-feces-containing liquid-suspended specimens. Furthermore, C T values for liquid-suspended specimens were 4 to 5 C T values lower (i.e., represented greater sensitivity) than those seen in direct swab analysis. Culture was equivalent to or better than Check-Direct CPE for 13/15 (87%) isolates tested in a limit-of-detection analysis. Detection of a carbapenemase gene at a C T cutoff value of <35 was culture confirmed in 23/24 (96%) of cases; however, C T values of >35 overlapped broadly between culture-positive (n ؍ 21) and culture-negative (n ؍ 36) specimens. Check-Direct CPE will likely prove most useful in high-prevalence areas or in outbreak settings where rapid carbapenemase detection is critical for infection control management. C arbapenemase-producing organism (CPO) infections are now globally epidemic, causing rising concern as they demonstrate easy transmissibility between patients and the environment, with increasing numbers of reported outbreaks (1-4). The morbidity and mortality associated with CPO infections are high, and effective therapeutic options are limited. Recent data show that bla KPC , bla NDM , and bla OXA-48 are the most common carbapenemases worldwide (5). Carbapenemase detection, however, can be complicated, and detection methods are not standardized among laboratories. Because gastrointestinal colonization with a CPO is a reservoir for transmission (6), routine CPO surveillance via collection of rectal/perirectal swabs or stool samples is increasingly being utilized in many infection control programs (7) and is recommended by public health agencies such as the Centers for Disease Control and Prevention (CDC) (8).Current methods for CPO surveillance generally rely on culture using selective media. The Clinical and Laboratory Standards Institute (CLSI) recommends performing carbapenemase testing on Enterobacteriaceae isolates (i) with imipenem or meropenem MICs of 2 to 4 g/ml or an ertapenem MIC of 2 g/ml (if using the 2010 M100-S20 brea...