Uncovering the predation habits of small arthropods such as mites can be a major challenge, as direct observations are not always feasible in natura. Therefore, PCR-based analyses of gut content, or body content when gut dissection is not applicable, are being increasingly used to detect remains of prey items. Here, we present an integrative approach to investigate predation of eggs of the viburnum leaf beetle Pyrrhalta viburni, an invasive pest in North America, by the oribatid mite species Trichoribates trimaculatus, to assess the potential of this mite as a biological control agent. We combined (1) manipulative laboratory experiments exposing beetle eggs to mites under controlled conditions and (2) body content molecular analyses using quantitative PCR (qPCR) to detect traces of consumed P. viburni egg DNA in mites. To account for risks of false-negative and false-positive results, mainly due to detectability problems, body surface contamination, and non-specific DNA amplification, we developed a stepwise procedure to analyze qPCR results. Egg integrity and survivorship were unaffected by mites in all experiments. However, traces of P. viburni DNA were detected in the body of mites exposed to P. viburni eggs, suggesting that they consumed either fragments of the chorion or fragments of the egg cap secreted by P. viburni females to protect the eggs, which was shown to contain P. viburni DNA. In conclusion, T. trimaculatus does not directly impact P. viburni eggs and should therefore not be considered as a biocontrol agent. The fact that mites did not directly predate eggs but contained traces of P. viburni DNA shows that detection of an organism's DNA within a presumed predator does not necessarily equal predation and that results of molecular analyses should be interpreted with caution in studies attempting to reconstruct trophic interactions.
1. The viburnum leaf beetle (VLB) is native to Eurasia and invasive in North America. It lays eggs inside young Viburnum twigs; eggs can be crushed by plant defences (wounding response) and attacked by natural enemies (parasitoids). 2. We evaluated plant defences and parasitism rates in 42 field sites in 11 European countries, on three host plants (V. opulus, V. lantana, and V. tinus), and under two main climatic conditions (Mediterranean, oceanic/continental). 3. Plant defences killed six times more eggs than natural enemies overall (25.4% vs. 4.1%). This effect was consistent for all host plants and climatic conditions. Egg survivorship and parasitism were lower in Mediterranean sites. 4. Parasitism was positively density-dependent, indicating a higher risk of parasitism on heavily infested twigs, but this effect was confounded with host plant species. 5. These results suggest that plant defences have been the dominant selection force driving the evolution of VLB oviposition behaviour.
French broom (Genista monspessulana) (Fabaceae) is a perennial species native to the Mediterranean basin. Introduced in the 19th century as an ornamental plant, it is currently invasive in California and Australia. The current research is focused on biocontrol with the use of the phytophagous weevil Lepidapion argentatum (Brentidae). Its capacity to develop both in the stem galls and pods of French broom makes it a promising candidate. The impact on the reproduction of French broom was studied in Southern France and revealed that it could effectively reduce the number of viable seeds by 18.8%, but also increased the number of aborted seeds by 10% within the attacked pods. To evaluate the specificity of L. argentatum, choice and no-choice tests were performed in 2012 and 2015 on a total of 36 non-target closely related species. Results revealed the presence of galls and larvae in the stems of seven species, including two endemic Californian lupines; i.e., Lupinus arboreus blue and Lupinus chamissonis. In the future, new tests will be conducted to determine if L. argentatum is able to complete its entire development lifecycle on the non-target species where galls have previously been observed.
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