Diagnostic markers for Planococcus ficus (Signoret) and Planococcus citri (Risso) by random amplification of polymorphic DNA‐polymerase chain reaction and species‐specific mitochondrial DNA primers

Abstract

Planococcus ficus (Signoret) and Planococcus citri (Risso) (Hom., Pseudococcidae) are important phytophagous components in different agroecosystems. The two species may coexist in the same environment and are most difficult to distinguish by morphological features. The aim of this study was to find genetic markers suitable for distinguishing P. ficus from P. citri, to assist in the rapid identification of field specimens. By using synthetic sex pheromone-baited traps, pure male populations of both species were collected from a vineyard and from a citrus orchard in northern Sardinia, Italy. Individual males of citrus and vine mealybugs were preliminarily examined by the random amplification of polymorphic DNA (RAPD) technique. Among twelve 10-mer random primers, the oligonucleotide OPL-12 generated several markers suitable for distinguishing between the two species. This primer was then used to characterize individual males and females of both mealybug species collected near pheromone-baited traps in vineyards and orange orchards from different geographic areas. Reference samples from other regions of southern Italy were also included. A clear differentiation of the two species was accomplished according to their pattern of amplification, thus confirming a high level of intra-specific genetic homogeneity. Consequently, two fragments of the cytochrome c oxidase I gene from P. citri and P. ficus were compared and two pairs of species-specific polymerase chain reaction (PCR) primers were developed based on diverging sequences. These primers allowed sensitive and reliable PCR identification of both males and females of P. citri and of P. ficus of different geographic origin.

Survey of mealybug (Hemiptera: Pseudococcidae) vectors of Ampelovirus and Vitivirus in vineyards of northwestern Italy

“…However, several mealybug species are very similar externally and their morphological identification, although accurate, requires a high level of expertise and hampers the recognition of nymphs and males (Millar 2002). These issues have recently solicited the use of DNA markers for the correct identification of the most agriculturally important species of pseudococcids (Cavalieri et al 2008;Demontis et al 2007;Saccaggi et al 2008). A further aim of this study was to develop a molecular identification tool for the species H. bohemicus.…”

Development of a Multiplex PCR for Identification of Vineyard Mealybugs

“…Multiplex PCR offers another rapid identiÞca-tion process for multiple species, where a number of species-speciÞc primers are used in a single PCR reaction. This process was successfully used by Demontis et al (2007) in Italy to separate Pl. ficus and Pl.…”

“…longispinus). Demontis et al (2007) and Cavalieri et al (2008) later described PCR-based separation of Pl. ficus and Pl.…”

Molecular diagnosis of two closely related mealybug species (Hemiptera: Pseudococcidae)

“…on the other hand, determination at the immature stages and males quite difficult by morphological method. This is why many molecular studies have recently been conducted to distinguish between closely related species (demontis et al, 2007; rung et al, 2008;Gullan et al, 2010; Malausa et al, 2011). Molecular studies can be used for different purposes when studying mealybugs (Hemiptera: Coccoidea: Pseudococcidae): dna barcoding, phylogeny, population genetics (demontis et al, 2007;Hardy et al, 2008; rung et al, 2008;Gullan et al, 2010;Malausa et al, 2010).…”