Phenacoccus madeirensis Green (Hemiptera: Pseudococcidae): New geographic records and rapid identification using a species-specific PCR assay

Abstract

The highly invasive Madeira mealybug Phenacoccus madeirensis Green (Hemiptera: Pseudococcidae) is a polyphagous pest which seriously harms crops of staple foods, fruits, vegetables, and ornamental plants. A method for rapid and accurate identification is needed to slow further spread of this pernicious pest. DNA barcoding and taxonomic keys were used to identify specimens of P. madeirensis collected in Rwanda and China. We designed a species-specific PCR (SS-PCR) primer pair for P. madeirensis based on variable sequences of the mitochondrial cytochrome c oxidase subunit I gene among P. madeirensis and 29 related species. This SS-PCR assay allows for rapid and precise identification. With this assay, no cross-reaction was detected from the 21 closely related mealybug species. All specimens from all ontogenic stages, adult debris, and individuals from 11 populations of P. madeirensis from different geographical areas and/or host plants were detected. The detection limit of this method was 192.58 ± 16.48 pg μL−1 of P. madeirensis DNA. These demonstrated the high sensitivity and effectiveness of this method. Using this method, we found the first described occurrence of P. madeirensis in Southern Province, Rwanda, indicating ongoing spread, and an entirely new invasion in Fujian, China. Invaded regions of China were delineated by a nationwide field study. The SS-PCR assay developed in this study is a rapid, straightforward, and reliable method for the identification of P. madeirensis, and will prove practical in plant quarantine, thus slowing or preventing further invasion and spread, and allowing for sustainable and effective management of this cosmopolitan invader.