The establishment of species-specific primers for the molecular identification of ten stored-product psocids based on ITS2 rDNA

Abstract

Psocids are important stored product pests found worldwide that can be spread through grain trade. Most stored-product psocids, including eggs, nymphs, and adults, are very small (~1 mm) and difficult to identify morphologically. Here, we collected 10 economically important stored-product Liposcelis spp. psocids (L. bostrychophila, L. entomophila, L. decolor, L. paeta, L. brunnea, L. corrodens, L. mendax, L. rufa, L. pearmani, and L. tricolor) from 35 geographical locations in 5 countries (China, Czech Republic, Denmark, Germany, and the United States). The ITS2 rDNA gene was extracted and sequenced. The interspecific genetic distance of the stored-product psocids was significantly higher than the intraspecific genetic distance according to the barcoding gap analysis. Ten pairs of species-specific primers based on the ITS2 rDNA were developed for psocid identification. The sensitivity estimation indicated that the species-specific primers could correctly amplify the target ITS2 gene and successfully identify psocids at 1.0 ng/mL. Additionally, these species-specific primers could quantify specificity and identify 10 stored-product psocids; this approach could also be used to accurately identify other stored-product psocids. This work provides a practical approach for the precise examination of 10 stored-product psocid species and also contributes to the development of an identification method using ITS2 rDNA.

Comparative Analysis of Mitochondrial Genomes among Twelve Sibling Species of the Genus Atkinsoniella Distant, 1908 (Hemiptera: Cicadellidae: Cicadellinae) and Phylogenetic Analysis

“…According to previous studies, the intraspecific divergences are rarely greater than 2% and most are less than 1% [ 66 ],. This was also verified in studies of several insect groups, such as Lepidoptera [ 67 ], Piophilidae [ 68 ], Blattidae [ 69 ], Liposcelididae [ 70 ], etc. The recent study of leafhoppers based on COX1 gene fragments also showed that the intraspecific divergence was less than 1% [ 71 ].…”

Rapid identification of the invasive fall armyworm Spodoptera frugiperda (Lepidoptera, Noctuidae) using species-specific primers in multiplex PCR

“…However, the application of species-specific primers based on ITS sequences in insect identification remains rare. A few studies of mosquitos, thrips, and stored-product psocids have been proposed using ITS species-specific primers 35,36,41,[43][44][45][46] . The results of the present study suggest that the ITS1 sequence variations of Spodoptera with high interspecific variations of > 16% and low intraspecific divergence of < 1.2% (Table 1) can be favourable markers with which to develop species-specific primers for FAW identification.…”

Molecular Identification of ten species of stored-product psocids through microarray method based on ITS2 rDNA

“…The extraction process is the same as that described by Zhao et al . in 2016 8 . The DNA was stored at −20 °C.…”

“…In order to overcome the limitations of morphological identification methods, many molecular technologies, including restriction fragment length polymorphism (PCR-RFLP) 3 , DNA barcoding 4 , species-specific primer PCR (SS-PCR) 7 , 8 , multiplex endpoint PCR and multiplex TaqMan qPCR 9 , have been developed for psocid identification. Molecular markers are the foundation for molecular identification, and both mtDNA (mitochondrial DNA) and rDNA (ribosomal DNA) are species-specific and can be used as molecular markers for species identification 10 , 11 .…”