Molecular Tools for the Detection and the Identification of Hymenoptera Parasitoids in Tortricid Fruit Pests.

Pierre Franck,Mariline Maalouly-Matar,Jérôme Olivares

doi:10.3390/ijms18102031

Pierre Franck, Mariline Maalouly-Matar + Show 1 more

Open Access

https://doi.org/10.3390/ijms18102031

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Abstract

Biological control requires specific tools for the accurate detection and identification of natural enemies in order to estimate variations in their abundance and their impact according to changes in environmental conditions or agricultural practices. Here, we developed two molecular methods of detection based on PCR-RFLP with universal primers and on PCR with specific primers to identify commonly occurring larval parasitoids of the tortricid fruit pests and to estimate parasitism in the codling moth. Both methods were designed based on DNA sequences of the COI mitochondrial gene for a range of parasitoids that emerged from Cydia pomonella and Grapholita molesta caterpillars (102 parasitoids; nine species) and a range of potential tortricid hosts (40 moths; five species) damaging fruits. The PCR-RFLP method (digestion by AluI of a 482 bp COI fragment) was very powerful to identify parasitoid adults and their hosts, but failed to detect parasitoid larvae within eggs or within young C. pomonella caterpillars. The PCR method based on specific primers amplified COI fragments of different lengths (131 to 463 bp) for Ascogaster quadridentata (Braconidae); Pristomerus vulnerator (Ichneumonidae); Trichomma enecator (Ichneumonidae); and Perilampus tristis (Perilampidae), and demonstrated a higher level of sensibility than the PCR-RFLP method. Molecular estimations of parasitism levels in a natural C. pomonella population with the specific primers did not differ from traditional estimations based on caterpillar rearing (about 60% parasitism in a non-treated apple orchard). These PCR-based techniques provide information about within-host parasitoid assemblage in the codling moth and preliminary results on the larval parasitism of major tortricid fruit pests.

Highlights

With the shift towards a reduced reliance on external inputs in agriculture, identifying management options that enhance pest control services has become a critical issue [1]
The use of individual primer sets to amplify uniquely sized fragments of the four most abundant C. pomonella parasitoids—A. quadridentata, P. vulnerator, P. tristis, and T. enecator—provided an accurate estimation of parasitism within immature Lepidoptera hosts; the molecular assessment of parasitism levels did not differ from estimates based on the monitoring of the emergences of parasitoid and moth adults, but was able to reveal the occurrence of multi-parasitism within the codling moth caterpillars and identify the species involved in this interaction, which is not possible with traditional rearing
The ten hymenopteran parasitoid species observed to emerge from C. pomonella and Grapholita molesta caterpillars collected in apple orchards from Western and Central Europe were all clearly differentiated according to their DNA barcodes

Summary

Introduction

With the shift towards a reduced reliance on external inputs in agriculture, identifying management options that enhance pest control services has become a critical issue [1]. The successful implementation of pest management programs requires a better understanding of the ecology behind the provision of ecosystem services [2] and methods to detect and identify the biodiversity linked with these services [3] Such methods are crucial when selecting biocontrol candidates [4] and when evaluating the efficiencies of biocontrol releases [5] or the impact of changes in agricultural practices [6]. PCR-amplified fragments can be subjected to RFLP, and sequenced or directly sized for species identification [3] These PCR-based techniques have been used to identify the presence of pests in contaminated food [11,12], to reveal prey in the gut or the faeces of predators [13], to determine the host from which a parasitoid adult emerged [14], or to detect parasite larvae inside their host [15]. They are at least as efficient as the traditional method of detection, which requires insect rearing during long periods or careful host dissection [17]

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