Géraldine Roux

Accurate and rapid identification of insect species is essential for effective biosurveillance, especially with increasing global trade and the spread of invasive pests. DNA barcoding has become a powerful tool for specimen identification, but its reliability depends heavily on the completeness and accuracy of reference databases. Many existing databases remain fragmentary and contain poorly annotated or misidentified records. Here, we present a curated DNA barcode reference library of 4097 DNA barcodes representing 169 species of longhorned beetles (Coleoptera: Cerambycidae), a group frequently intercepted in European biosurveillance programmes using multi‐pheromone traps. By prioritizing taxonomic validation and data quality, this library provides a robust resource for accurate specimen identification. Beyond biosurveillance, it also supports ecological research, as cerambycids are valuable bioindicators of forest health. Our results highlight the critical role of expert curation in ensuring the accuracy of barcode libraries used for both pest monitoring and biodiversity assessment.

Monitoring regulated quarantine plant pathogens and, when relevant, their vectors is compulsory in the European Union. Local Monochamus species (Coleoptera, Cerambycidae) vector the pine wood nematode, Bursaphelenchus xylophilus , a non-native pest already established in most of Portugal. Only 29 M. galloprovincialis individuals were trapped in Belgium in ten years (2013–2022), despite a dense coverage of pheromone-baited traps, suggesting absence or a very rare local occurrence in the country. In the northern neighbouring countries, only one single established population is known in The Netherlands and one in Denmark. A species distribution model based on pheromone-trap catches (negative and positive) of M. galloprovincialis from 4,914 traps in 29 European countries between 2008 and 2019 was developed, using the overall climate conditions and the distribution of seven pine tree species as explanatory variables. The effect of spatial scale was tested with a multi-scale approach. With a 225*225 km spatial grain, the major explanatory variables were the mean diurnal temperature range and, to a lesser extent, the presence of Pinus spp. The model predicted a low probability of presence in Belgium, the Netherlands, Great Britain and north-western Germany compared to southern Europe. Genotyping allowed to conclude that at least some of the beetles caught in Belgium originated from foreign locations. All catches were located close to entry points, suggesting introduction with imported material. The small size of most of the Belgian pine stands may also explain the absence or apparently transient status, or rareness of Monochamus spp. This study thus suggests that surveys in Belgium should privilege entry points rather than local forest stands.

In the Western Palaearctic, the congeneric bark beetles Hylurgus ligniperda and Hylurgus micklitzi are developing on pine trees. Although H. ligniperda has a larger native range in Europe, both species often occur in sympatry in the Mediterranean Basin. They share rather similar life histories, thus providing an excellent model for comparative studies on species differentiation and ecological traits. Moreover, along with increasing human activities and international trade, H. ligniperda has invaded all continents, whereas H. micklitzi remains confined to the Mediterranean Basin. Our research utilised COI markers to assess the genetic diversity and structure, alongside the demographic history, of both species and revealed marked differences. H. micklitzi exhibited low genetic diversity and shallow population structures with restricted expansion. In contrast, H. ligniperda displayed a longer and more complex post-glacial evolutionary route with four distinct clades originating from separate glacial refugia, coupled with considerable genetic variability across Europe. Comparative analyses about the two species suggest that species traits, such as larger body size, increased voltinism, broader ecological niches and frequent introduction events may have helped the fast global invasion and spread of H. ligniperda over the past century.

The Scots pine cone weevil, Pissodes validirostris Gyll. (Coleoptera: Curculionidae) widely distributed from Europe, across to North-Eastern China, consumes the cones of Mediterranean pines, as well those of the sylvestris section. The use of multiple host pine species with limited and patchy distributions, combined with low dispersal rates, likely contributed to significant genetic and morphological variation among P. validirostris populations in its native range. P. validirostris is being considered as a biological control agent for wilding pines in the southern hemisphere and for this programme to proceed, elucidating its intraspecific variability is necessary. This study aimed to explore the phylogeography of P. validirostris in its native range in Europe using mitochondrial and nuclear genetic markers as well as morphological characters. The integrative taxonomic analysis revealed high genetic structuring in P. validirostris populations, identifying three main phylogeographic clades: Clade A (Iberian Peninsula, associated with Pinus pinaster and Pinus halepensis), Clade B (Central Europe), and Clade C (Northern and Eastern Europe), both associated with Pinus species of the sylvestris section. The morphometric data showed that adults of the Iberic Peninsula clade were significantly larger than those of the two others. When compared to other Nearctic and Palearctic congeneric species, P. validirostris formed a monophyletic group, likely consisting of different evolutionary lineages within the sampled distribution. The findings highlight the necessity of genetic confirmation for insect taxonomy across wide distributions or multiple host trees. Clarifying whether host specificity or geography drives P. validirostris population structuring will facilitate the selection of a specific clade for wilding pine biological control.

Individual sorting and identification of thousands of insects collected in mass trapping biosurveillance programmes is a labour-intensive and time-consuming process. Metabarcoding allows the simultaneous identification of multiple individuals in a single mixed sample and has the potential to expedite this process. However, detecting all the species present in a bulk sample can be challenging, especially when under-represented non-native specimens are intercepted. In this study, we quantified the effectiveness of DNA metabarcoding at detecting exotic species within six different mock communities of native and non-native species of European xylophagous cerambycid beetles. The main objective is to compare three different sequencing technologies (MinION, Illumina and IonTorrent) to evaluate which one is the most suitable in this context. Additionally, dry and wet (monopropylene glycol and water) collection methods were compared. Although not observing significant differences in the total number of species detected amongst the three sequencing technologies, the MinION detected a greater number of species in field-like samples. All three sequencing technologies achieved success in detecting and identifying closely-related species and species in low abundance. The capture method of insects in the field greatly influenced sample preservation and detection. Individuals captured in traps containing monopropylene and water had lower DNA concentration, leading to lower species detection rates compared to individuals killed using just an insecticide without any collection medium.

Individual sorting and identification of thousands of insects collected in mass trapping biosurveillance programs is a labour intensive and time-consuming process. Metabarcoding, allows for the simultaneous identification of multiple individuals in a single mixed sample and has the potential to expedite this process. However, detecting all the species present in a bulk sample can be challenging, especially when under-represented non-native specimens are intercepted. In this study, we quantified the effectiveness of DNA metabarcoding at detecting exotic species within six different mock communities of native and non-native species of European xylophagous cerambycid beetles. The main objective is to compare three different sequencing technologies (MinION, Illumina, and IonTorrent) to evaluate which one is the most suitable in this context. Although we did not observe significant differences in the total number of species detected between the three sequencing technologies, MinION detected a greater number of species on field-like samples. All three sequencing technologies achieved in detecting and identifying closely related species and species at low abundance. The capture method of insects in the field greatly influences sample preservation and detection. Individuals captured in traps containing monopropylene and water had lower DNA concentration, leading to lower species detection rates compared to individuals killed using just an insecticide without any collection medium.

Key message: The invasive pine wood nematode is a major threat to pine forests worldwide, causing extensive tree mortality. Although scientific knowledge and control measures are continuously improving, important gaps remain. We argue that some key questions, notably related to early detection and pest management, need to be urgently tackled in countries at risk of invasion such as France.

Individual sorting and identification of thousands of insects collected in mass trapping biosurveillance programs is a labor intensive and time-consuming process. Metabarcoding, which allows for the simultaneous identification of multiple individuals in a single mixed sample, has the potential to expedite this process. However, detecting all the species present in a bulk sample can be challenging, especially when under-represented non-native specimens were intercepted. In this study, we quantified the effectiveness of metabarcoding at detecting exotic species within six different mock communities including or not native and non-native species of European xylophagous cerambycid beetles. Although we did not observe significant differences in the total number of species detected between MinION, Illumina, and IonTorrent sequencing technologies, a greater number of individuals was detected and identified to species using MinION, including the detection of three non-native cerambycids. The three sequencing technologies also showed similar results in detecting and identifying closely related species and species at low abundance. The capture method appears to greatly influence sample preservation and detection. Indeed, individuals captured in traps containing monopropylene and water had both lower DNA concentration leading to lower species detection rates compared to individuals killed using just an insecticide without any collection medium.

Individual sorting and identification of thousands of insects collected in mass trapping biosurveillance programs is a labor intensive and time-consuming process. Metabarcoding, which allows for the simultaneous identification of multiple individuals in a single mixed sample, has the potential to expedite this process. However, detecting all the species present in a bulk sample can be challenging. In this study, we quantified the effectiveness of metabarcoding at detecting all species in six different mock communities of xylophagous cerambycid beetles. No significant differences in the number of species detected were observed between MinION, Illumina, and IonTorrent sequencing technologies. However, a greater number of individuals was detected and identified to species using MinION. In addition, the proportion of reads assigned to the species level was higher with Illumina technology. The three sequencing technologies also showed similar results in detecting and identifying closely related species and species at low abundance. The capture method greatly influences sample preservation and detection. Indeed, individuals captured using monopropylene and water had both lower DNA concentration and species detection rates compared to individuals killed using just an insecticide without any collection medium.