| Summary: | Monitoring of <i>Pentastiridius leporinus</i> (Hemiptera: Auchenorrhyncha: Cixiidae), representing the main vector of the syndrome ‘basses richesses’ (SBR) disease in sugar beet is based on morphological identification. However, two other cixiid species, <i>Reptalus quinquecostatus</i> and <i>Hyalesthes obsoletus</i> with similar external characters are known to appear in sugar beet fields and are challenging to be distinguished from <i>P. leporinus</i>. We present a PCR-based method for species-specific detection of both male and female <i>P. leporinus</i>, directly after sweep net collection or after up to 18 months long term storage on sticky traps. Two methods of DNA template preparation, based on a commercial extraction kit or on simple grinding in phosphate-buffered saline (PBS) were compared. The latter method was also established for eggs and all five nymphal instars of <i>P. leporinus</i> from a rearing. Furthermore, in silico primer analysis showed that all Auchenorrhyncha species including far related species reported from sugar beet fields can be differentiated from <i>P. leporinus</i>. This was PCR-confirmed for the most common Auchenorrhyncha species from different German sugar beet fields. Sequence analysis of the <i>P. leporinus</i> mitochondrial cytochrome oxidase I gene (<i>COI</i>) amplicon showed a close relationship to <i>COI</i> from <i>P. beieri</i> but separated from the <i>Reptalus</i> and <i>Hyalesthes</i> species which are grouped into the same family Cixiidae. We present a sensitive, cost- and time-saving PCR-based method for reliable and specific detection of eggs and all nymphal instars, as well as male and female <i>P. leporinus</i>, after different methods of planthopper collection and template DNA template preparation that can be used in large scale monitoring assays.
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