Drosophila suzukii (Matsumura) (Diptera: Drosophilidae) is a ubiquitous global pest of several fruit crops. Trapped adult numbers are used to monitor populations and make control decisions, but differentiating D. suzukii from other trapped Drosophila spp. is laborious. We developed a real-time PCR method for specific detection and semi-quantification of D. suzukii from trap samples. The PCR assay did not amplify DNA from 29 other Drosophilidae species tested. Drosophila suzukii was detected from ≥0.96 pg target DNA and from laboratory samples containing one D. suzukii in 2000 other Drosophila spp. flies. We tested DNA stability of one D. suzukii in 100 Drosophila spp. flies in water or ethanol at 20, 25, or 30°C for 1, 4, or 7 d. Only water at 30°C for 7 d fully impaired D. suzukii DNA detectability. Substituting mouthwash for water resulted in D. suzukii detection in all samples held for 7 d at 30°C or daily fluctuating temperatures of 33/23°C. Traps with mouthwash as a drowning liquid had D. suzukii captures equal to traps with water. A calibration curve was established using samples in mouthwash containing 1/1,000—100/1,000 D. suzukii/total Drosophila spp. flies and incubated at 25°C for 7 d. The curve had a coefficient of determination (R²) of 0.9279 between D. suzukii numbers from the PCR and the true D. suzukii numbers in samples prepared in 70% ethanol. Collecting samples in mouthwash is expected to improve detection accuracy, and the qPCR method can be a useful tool to support D. suzukii monitoring and management.