First records of <i>Ceroplastes ceriferus</i> (Fabricius) (Hemiptera: Coccidae) and <i>Ceroplastes japonicus</i> (Gray) in Switzerland identified by DNA barcoding

Wax soft scales (Hemiptera: Coccidae) include many important pest species of agricultural and ornamental plants which spread along international trading networks plant material. Here we report the first findings wax Ceroplastes japonicus ceriferus for Switzerland. The two new were collected on Laurus nobilis Vaccinium myrtillus, respectively, identified by DNA barcoding. Les cochenilles lécanides comprennent de nombreuses espèces d’organismes nuisibles d’importance sur les plantes agricoles et ornementales se disséminent via réseaux internationaux commerce matériel végétal. premiers signalements en Suisse des sont présentés dans cet article. spécimens ont été collectés respectivement myrtillus identifiés par barcoding moléculaire. Boc?o??e ?o??o???o??? ?????a?? ??o??e ?a???e ???? op?a????o?, ?pe???e ??? ce??c?oxo???c??e???x ? ?e?opa?????x pac?e??? pac?poc?pa?????ec? ?o ?e??y?apo???? ce??? ?op?o??? pac?e????? pac???e????? ?a?ep?a?o?. ??ec? ?? coo??ae? o ?ep??x o??apy?e???x ?oc?o??x ?o??o???o???o? ? ??e??ap??. ??? ??e ?o??e ?axo??? ???? co?pa?? ?a coo??e?c??e??o ??e???????po?a?? c ?o?o??? ?H?-??p?x?o??po?a???. unintended dispersal invasive insect pests global trade represents an increasing threat horticultural production that can cause serious economic damage (Bacon al., 2012; Ali 2020). These insects from family Coccidae (Miller & Miller, 2003), has more than 1200 described to date (García Morales 2016). subfamily Ceroplastinae is characterized a thick waxy layer covering entire (Qin Gullan, 1994). One genus within this Gray, contains have been introduced EPPO region Global Database six with confirmed records Europe. Of all members Ceroplastes, only rusci regulated in EU Switzerland, classified as non-quarantine pest. Although Papadopoulou al. (2020) provide very helpful key morphological identification occurring Europe, their classical taxonomy remains challenging. Accurate fast prerequisite research disciplines factor implementation stringent management strategies against (Floyd 2010; Hodgetts Traditional based characteristics often time-consuming relies expert knowledge skilled taxonomic specialists (Hebert 2003; be especially challenging they are found immature stages lacking distinct characters (Hodgetts 2016; Saccaggi Blaser 2018). Further problems arise if specimens damaged or no available To circumvent these drawbacks, alternatively performed using molecular methods such antibody-based, protein-based genetic-based approaches (Armstrong Ball, 2005; Among them, powerful generic tool wide range easily standardized between different laboratories 2003). PCR amplification sequencing conserved signature sequence subsequently queried database (e.g. Bold, GenBank Q-bank) containing reference sequences previously (Sujeevan Hebert, 2007; Benson 2008; Floyd Bonants 2013; it advantages, limited fact method identify well primer information limitations when degree genetic differentiation fragment too low enable clear closely related 2005). In implemented regular import control process consignments standard potential quarantine (Blaser addition its use diagnostic purposes, combined next-generation (NGS) was evaluated promising biodiversity monitoring schemes (Gueuning 2019). Studying diversity COI among (C. floridensis, C. japonicus, ceriferus, pseudoceriferus, rubens kunmingensis) demonstrated properly resolves taxa into reciprocally monophyletic clades congruent units derived characters. aim present study reliably confirming record Canton Valais March 2021. finding not official survey activity, but project collaboration farmer. Ticino private person December 2020. Specimens both collection sites preserved 70% ethanol prior extraction. A map displaying generated QGIS version 3.10.8 free geodata Swiss Federal Office Topography, swisstopo. extracted approximately 1 mm3 tissue one adult specimen each following protocol modified Kawasaki (1990), Brunner (2002) Frey (1995). brief, placed 1.2-mL microtubes (Qiagen AG, Hilden, Germany), 2-mm diameter steel bead (Retsch GmbH, Haan, Germany) 100 ?L lysis buffer (10 mM Tris-HCL pH 8.0, EDTA, 0.5% Tween 20, ?g/mL Proteinase K). After grinding samples twice 3 min at 30 Hz mixer mill MM 300 GmbH), homogenization product incubated 95? thermomixer comfort (Eppendorf Hamburg, Germany). amplify barcode fragments 5? end mitochondrial gene cytochrome oxidase subunit (COI), polymerase chain reaction (PCR) run thermal cycler (Senso-Quest Göttingen, For sample Valais, amplified primers C1-1554F (5?-CAGGAATAATAGGAACATCAATAAG-3?) C1-2342R (5?-ATCAATGTCTAATCCGATAGTAAATA-3?). Ticino, assembled overlapping consensus combination above together C1-J-2183 (5?-CAACATTTATTTTGATTTTTTGG-3?) C1-N-2568 (5?-GCWACWACRTAATAKGTATCATG-3?) (Brady 2000). carried out total volume 20 extract, 0.4 ?M × HotStarTaq cycling conditions: 15 95?, followed 35 cycles 40 s 45?, ramping over 60 60? 2 72?. final elongation step 7 72?, cleaned NucleoFast 96 system (Marcherey-Nagel Düren, manufacturer's subsequent linear 8 diluted 1:10 grade water, 0.2 either forward reverse (see above) BigDye Terminator v1.1 Ready Reaction Mix (Applied Biosystems) (Waltham-Massachusetts-United States). removing unincorporated dye terminators DyeEx Kit AG) according protocol, sequenced SeqStudio Genetic Analyzer Biosystems). Applying de-novo assembly function Geneious 10.0.9 assembler default settings, blasted publicly accessible databases Bold Kearse 2012). Classification best matches Max Score values BLAST results. canton uploaded accession numbers MZ836057 MZ836034, respectively (Benson 2008). plastic tunnel 0.1 ha. Approximatively 5% heavily infested. garden single plant, where other infested (Figures 2). novo step, trimmed 654 867 bp retained cantons respectively. When querying databases, clearly (GenBank, identity 98.17%, coverage 100%; similarity 100%). second-best match pseudoceriferus substantially lower percentage 95.41%, 95.7%). Querying sample, 99.85%, 75%; 99.27%). Additionally considerably assigned kunmingensis 92.23%, 65%; 97.07%). locations Switzerland shown Figure 3. represent recently initiated person, another unrelated project, stressing occasional vigilant observers detection. This shows importance awareness raising. demonstrate necessity efforts form systematic surveys conducted national level early detection foreign pathogens. paper further emphasizes cryptic species. Particularly cases group scarce, tools may pillar sampling efforts. Open Access Funding provided Agroscope. [Correction added 16 May 2022, after online publication: CSAL funding statement added.]