Assessment of the diversity of Aphthona flea beetles for biological control of leafy spurge on the prairies

Leafy spurge (Euphorbia esula) is an herbaceous, perennial weed of European origin. It is toxic to cattle and invasive on rangelands in North America. Beginning in the 1980s, six European species of Aphthona flea beetles (Chrysomelidae) were released in North America to control this weed. At sites where control has been achieved, success has been attributed mainly to A. nigriscutis and A. lacertosa. In the present study, we collected Aphthona at historical release sites to re-assess Aphthona diversity in western Canada. Specimens were identified to species using colour, external morphology, and the shape of the genitalia. For a subset of these specimens, plus pinned voucher specimens, we obtained DNA sequences for COI, ITS2, and 28S genes. The voucher specimens were collected in Europe in the 1980s and identified at that time using morphological traits. COI sequences identified five clades corresponding to five of the Aphthona species originally released in North America. However, specimens identified as A. czwalinae grouped with A. nigriscutis; A. flava and A. cyparissiae co-occurred in different clades. Within the A. lacertosa clade, there was evidence of three subspecies or possibly a species complex. ITS2 and 28S sequences each identified only two clades, which corresponded to ‘brown’ beetles (A. cyparissiae, A. flava, A. nigriscutis) and ‘black’ beetles (A. lacertosa). ITS2 and 28S sequences for A. czwalinae were unavailable. Aphthona flava was absent in field collections. Based on these collective results, adoption of genetic-based markers is recommended in future studies of this genus to verify species determinations.