【Background】Paraleyrodes pseudonaranjae Martin, a new invasive species in southern China, is a worldwide pest, causing serious damage to a wide range of host plants. Further spread of this whitefly species in China is expected. Most of whitefly species are small in size and similar in morphology, which makes them hard to be identified quickly. 【Method】In the present study, we developed DNA markers to rapidly identify P.pseudonaranjae. The fragments of mitochondrial DNA cytochrome oxidase Ⅰ (mtDNA COⅠ) gene of P.pseudonaranjae and six other whitefly species were amplified (using COⅠ gene 5′ end universal primers LCO-1490/HCO-2198) and sequenced. A pair of P.pseudonaranjae speciesspecific COⅠ (SSCOⅠ) primers (PPZYF1/PPZYR1) were developed. The target fragment amplified by the SS-COⅠ primers PPZYF1/PPZYR1 was 233 bp long. 【Result 】The species specificity of the primer pair was validated using eight other whitefly species, including different cryptic species of Bemisia tabaci (Gennadius) (MEAM1, MED, Asia Ⅱ 3 and Asia Ⅱ 1) as well as Trialeurodes vaporariorum (Westwood), Dialeurodes citri (Ashmead), Aleurocanthus spiniferus (Quaintance) and Aleurodicus dispersus (Russell), all of them common in China. All P.pseudonaranjae specimens were detected, and no cross reactions with other whitefly species were obtained. The method was tested on single male and female adult, single redeyed nymph, single 2nd-and 3rd-instar nymph, and even single egg and newly emerged nymph, and proved to be applicable for all of these life stages. When the dilution was 1/40960 of a whole female adult of P.pseudonaranjae, the 233 bp DNA fragment could be identified for all repeats. 【Conclusion and significance】The technique would be useful in quarantine at ports of entry and in pest detection and monitoring during transportation of ornamental plants, fruits and seedlings.