Reliable molecular detection of small hive beetles

Invasive species require adequate reliable detection methods to mitigate their further spread and impact. However, the reliability of molecular detection methods is often hampered by both false positives (Error type I) and false negatives (Error type II). At present, the reliability of the four published molecular detection methods for small hive beetles (SHB), Aethina tumida, has not been rigorously evaluated considering their extensive genetic diversity. Here, we performed intra- and interlaboratory comparisons of the four available methods using SHB samples representing 78 regions from 27 countries on five continents, beetles from the same genus (Aethina concolor, A. inconspicua, A. flavicollis and A. major), as well as western honey bees, Apis mellifera, and ectoparasitic mites Varroa destructor. The data show that the Idrissou et al. (2018) and Li et al. (2018) methods avoid both false positives and false negatives probably due to lower sensitivity to nucleotide mismatches on the primer and probe’s target sequences. Further, the Li et al. (2018) method can be considered more sensitive because the fluorescent amplification curve crosses the threshold at lower Cq values compared to the Idrissou et al. (2018) one. In light of our data, the Li et al. (2018) method is the most reliable molecular diagnostic tool for SHB. We therefore recommend using this method as it will contribute to management efforts of this invasive species.