A diagnostic real-time PCR assay for the rapid identification of the tomato-potato psyllid, Bactericera cockerelli (Šulc, 1909) and development of a psyllid barcoding database

The accurate and rapid identification of many insect pests is an important step in the prevention and control of outbreaks in areas that are otherwise pest free. The potato-tomato psyllid Bactericera cockerelli (Šulc, 1909) is the main vector of ‘ Candidatus Liberibacter solanacearum’ on potato and tomato crops in Central and Northern America and New Zealand. This study describes the design and validation of the first species - specific TaqMan probe-based real-time PCR assay, targeting the ITS2 gene region of B. cockerelli . The assay successfully detected B. cockerelli genomic DNA from adults (100% accuracy, n=72); immatures (100% accuracy, n=26) and eggs (100% accuracy, n=25) . This assay also detected DNA from cloned plasmids containing the ITS2 region of B. cockerelli (100% accuracy, n=24). The assay showed 0% false positives when tested on genomic and cloned DNA from 73 other psyllid species collected from across Europe, New Zealand and Mexico. This included 8 other species in the Bactericera genus and the main vectors of ‘ Candidatus Liberibacter solanacearum’ worldwide. The limit of detection for this assay at optimum conditions was 0.000001ng DNA (~200 copies) of ITS2 DNA which equates to around a 1:10000 dilution of DNA from one single adult specimen. This assay is the first real-time PCR based method for accurate, robust, sensitive and specific identification of B. cockerelli from all life stages. It can be used as a surveillance and monitoring tool to further study this important crop pest and to aid the prevention of outbreaks, or to prevent their spread after establishment in new areas.