Simple, rapid, and sensitive screening methods are the key to prevent and control the spread of foodborne diseases. In this study, a simple visual colorimetric assay using magnetic nanoparticles (MNPs) and gold nanoparticles (AuNPs) was developed for the detection of Vibrio parahaemolyticus. First, the aptamer responding to V. parahaemolyticus was conjugated onto the surface of MNPs and used as a specific magnetic separator. In addition, the aptamer was also immobilized on the surface of AuNPs and used as a colorimetric detector. In the presence of V. parahaemolyticus, a sandwich structure of MNP-aptamer-bacteria-aptamer-AuNPs is formed through specific recognition of the aptamer and V. parahaemolyticus. The magnetic separation technique was then applied to generate a detection signal. Owing to the optical properties of AuNPs, a visual signal could be observed, resulting in an instrument-free colorimetric detection. Under optimal conditions, this assay shows a linear response toward V. parahaemolyticus concentration through the range of 10-106 cfu/mL, with a limit of detection of 2.4 cfu/mL. This method was also successfully applied for V. parahaemolyticus detection in spiked raw shrimp samples.