Riemerella anatipestifer (RA) infections cause major economic losses in the duck industry. Detection of RA using conventional assays is time-consuming and laborious. In this study, a simple and rapid assay for the detection of RA was established based on the GroEL gene sequence of RA using loop-mediated isothermal amplification (LAMP) with a set of six primers (two outer primers, two inner primers, and two loop primers). This assay was able to detect all the tested RA strains with different serotypes. A minimum of 10 colony-forming units (CFU) of RA was detected, which represents 50-fold higher sensitivity than that of the standard polymerase chain reaction (PCR) method. This assay showed good specificity to RA strains and did not react with any other species of bacteria. The assay is rapidly completed and the amplification is achieved at a minimum of 20 min at 65 C. Furthermore, the assay successfully detected RA in the liver samples of ducklings infected with RA, suggesting that the assay could be used for the clinical diagnosis of RA infection.