[Objectives]  Fruitflies (Diptera, Drosophilidae) are important globally distributed quarantinable pests. They generally lay eggs in fruits and vegetables, and their larvae feed on these after hatching, causing serious economic loss to fruit and vegetable growers. The eggs, larvae, and pupae of fruitflies are frequently discovered in quarantine ports, however, these life-stages are difficult to identify because fruitfly species are traditionally distinguished on the basis of adult morphology. Morphological identification of eggs, larvae, and pupae therefore requires these to be reared to the adult stage, which takes a long time. It is therefore useful to develop rapid and accurate molecular methods that can identify fruitfly eggs, larvae, and pupae. [Methods]  Species specific, simple sequence repeats (SSR) were discovered in the genomes of four quarantined fruit fly species; Ceratitis capitate, Bactrocera cucurbitae, Bactrocera dorsalis and Bactrocera tryoni, using a well-designed bioinformatics pipeline. Species-specific primers were designed from these SSRs and screened using genomic DNA extracted from specimens of different fruitfly species as templates. The polymerase chain reaction (PCR) was then used to amplify and detect SSRs in DNA samples obtained from fruitfly eggs, larvae and pupae. [Results]  Twenty species-specific SSRs were obtained from the four quarantined fruitfly species. Three species-specific primers were designed which produced single bands of 1 251, 1 307 and 823 bp, respectively and were confirmed to reliably distinguish C. capitata, B. cucurbitae and B. dorsalis. [Conclusion]  Bioinformatics analysis of genomic sequences allows us to identify species-specific SSR. After the optimization of species-specific primers designed from these SSR sequences, PCR can be used to accurately identify fruitflies. This work provides a theoretical basis and technical support for inspectors to reliably identify fruitfly eggs, larvae and pupae intercepted at ports.