Identification of Phenacoccus solenopsis Tinsley (Hemiptera: Pseudococcidae) with species-specific COI (SS-COI) primers.
Phenacoccus solenopsis Tinsley, a new invasive species in China, is a worldwide pest causing serious threat to the production of agriculture and forestry. Morphological identification of this mealybug species is limited by high degree of similarity and polymorphism. In the present study, a method was described for the development of DNA markers for rapidly identifying P. solenopsis. A pair of universal primers was designed based on published mitochondrial DNA cytochrome c oxidase subunit I (mtDNA COI) gene sequences of mealybugs in GenBank. The mtDNA COI genes from P. solenopsis and seven other mealybug species common in China including Pseudococcus comstocki Kuwana, Planococcus lilacius Cockerell, Maconellicoccus hirsutus (Green), Saccharicoccus sacchari (Cockerell), Dysmicoccus neobrevipes Beardsley, Planococcus minor Maskel and Phenacoccus solani Ferris were amplified and sequenced. And then one pair of species-specific COI (SS-COI) primers was designed. The SS-COI primers amplified a single band of 546 bp from P. solenopsis. The specificity of the primer pair was validated using the seven other mealybug species mentioned above. The results showed that all and only P. solenopsis specimens were detected, and no cross reactions with other mealybugs were observed. The method was tested with individuals of the egg, 1st, 2nd and 3rd instar nymph and female adult, and demonstrated to be applicable for all life stages. Furthermore, the primer set was tested with one P. solenopsis population from Pakistan and fourteen P. solenopsis populations from invaded areas in China and proved to be applicable for all geographic populations. The diagnostic PCR assay developed here provides a quick, simple and reliable molecular technique for the identification and monitoring of P. solenopsis, which will be useful in intercepting and blocking the further spreading of P. solenopsis.